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2016 09-16-fairdom

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Keynote talk given at Fairdom User meeting http://fair-dom.org/communities/users/barcelona-2016-first-user-meeting/ .

I begin by summarising how we apply molecular approaches to understand social behaviour in ants. Subsequently, I give an overview of the data-handling challenges the genomic bioinformatics community faces. Finally, I give an overview of some of the tools and approaches my lab have developed to help us get things done better, faster, more reliably and more reproducibly.

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2016 09-16-fairdom

  1. 1. A major transition in social evolution & some data tidbits y.wurm@qmul.ac.uk https://wurmlab.github.io
  2. 2. © Alex Wild & others
  3. 3. © National Geographic Atta leaf-cutter ants
  4. 4. © National Geographic Atta leaf-cutter ants
  5. 5. © National Geographic Atta leaf-cutter ants
  6. 6. Oecophylla Weaver ants © ameisenforum.de
  7. 7. © ameisenforum.de Fourmis tisserandes
  8. 8. © ameisenforum.de Oecophylla Weaver ants
  9. 9. © forestryimages.org© wynnie@flickr
  10. 10. Tofilski et al 2008 Forelius pusillus
  11. 11. Tofilski et al 2008 Forelius pusillus hides the nest entrance at night
  12. 12. Tofilski et al 2008 Forelius pusillus hides the nest entrance at night
  13. 13. Tofilski et al 2008 Forelius pusillus hides the nest entrance at night
  14. 14. Tofilski et al 2008 Forelius pusillus hides the nest entrance at night
  15. 15. Avant Workers staying outside die « preventive self-sacrifice » Tofilski et al 2008 Forelius pusillus hides the nest entrance at night
  16. 16. Dorylus driver ants: ants with no home © BBC
  17. 17. Animal biomass (Brazilian rainforest) from Fittkau & Klinge 1973 Other insects Amphibians Reptiles Birds Mammals Earthworms Spiders Soil fauna excluding earthworms, ants & termites Ants & termites
  18. 18. Well-studied: • behavior • morphology • evolutionary context
  19. 19. Well-studied: • behavior • morphology • evolutionary context • ecology Genetic basis?
  20. 20. REPORTS onMarch12,2013www.sciencemag.orgDownloadedfrom Solenopsis invicta fire ants are a big problem! verywellstudied! Ascunceetal2011
  21. 21. Solenopsis invicta fire ant: two social forms •1 large queen •Independent founding •Highly territorial •Many sizes of workers •2-100 smaller queens •Dependent founding •No inter-colony aggression •All workers similar size Single-queen form: Multiple-queen form:
  22. 22. Fire ants + Population genetics:Allozyme screen Ken Ross “starch gel”+ 1 2 3 L. Keller
  23. 23. Allozyme screen Social form associated to Gp-9 locus Frequency of the most common allele Locus! 0.3! 0.4! 0.5! 0.6! 0.7! 0.8! 0.9! 1.0! Single queen!Multiple queen! Est-6!Est-4! G 3pdh-1!C a-4!Pgm -4!Ddh-1!Pro-5! Pgm -3!Acoh-5!acoh-1!A cy-1!Pgm -1!Aat-2!Gp-9! Ken Ross and colleagues Laurent Keller and colleagues
  24. 24. Single queen form Multiple queen form Ken Ross and colleagues Laurent Keller and colleagues Social form completely associated to Gp-9 locus
  25. 25. bbbbBB BB Bb bb Ken Ross and colleagues Laurent Keller and colleagues Single queen form Multiple queen form Social form completely associated to Gp-9 locus (>15% )(< 5% )
  26. 26. bbBB BB Bb x Gp-9 bb females rare Ken Ross and colleagues Laurent Keller and colleagues Single queen form Multiple queen form Social form completely associated to Gp-9 locus (>15% )(< 5% )
  27. 27. BB BB Bb Ken Ross and colleagues Laurent Keller and colleagues Single queen form Multiple queen form Social form completely associated to Gp-9 locus (>15% )(< 5% )
  28. 28. BB BB Bb x Ken Ross and colleagues Laurent Keller and colleagues Single queen form Multiple queen form Social form completely associated to Gp-9 locus (>15% )(< 5% )
  29. 29. BB BB Bb x x Ken Ross and colleagues Laurent Keller and colleagues Social form completely associated to Gp-9 locus Single queen form Multiple queen form (>15% )(< 5% )
  30. 30. BB BB Bb x x x Ken Ross and colleagues Laurent Keller and colleagues Single queen form Multiple queen form (>15% )(< 5% ) Social form completely associated to Gp-9 locus
  31. 31. • Is this gene the single überregulator? Social form completely associated to Gp-9 locus
  32. 32. • Is this gene the single überregulator? maybe 1/14th of the genome? •Only 14 allozyme markers Locus! 0.3! 0.4! 0.5! 0.6! 0.7! 0.8! 0.9! 1.0! Single queen!Multiple queen! Est-6!Est-4! G 3pdh-1!C a-4!Pgm -4!Ddh-1!Pro-5! Pgm -3!Acoh-5!acoh-1!A cy-1!Pgm -1!Aat-2!Gp-9! Social form completely associated to Gp-9 locus
  33. 33. This changes everything. Any lab can sequence anything! http://genome.gov/sequencingcosts
  34. 34. Bb unfertilised eggs haploid ♂ Gp-9 B Gp-9 b Gp-9 B Gp-9 b Gp-9 b Gp-9 B 38 B♂ & 38 b♂ RAD genotyping
  35. 35. Identify polymorphism individual x locus genotype table RAD genotyping: sequencing the same 0.01% of the genome in many individuals A B C D E F L1 A C A A C C L2 G G T - T G L3 - A G A - G L4 C - - G G C L5 T T C T C - L6 G A A - - G 2419loci 38 B & 38 b 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20+ Amount of variance explained per principal component %VarianceExplained 051015202530 12.7% 6.1% 5.4% 4.8% 4.7% 3.9% 3.5% 3.2% 3.1% 2.9% 2.8% 2.6% 2.4% 2.3% 2.2% 2.0% 1.9% 1.7% 1.6% 30.2% 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20+ Amount of variance explained per principal component %VarianceExplained 051015202530 PCA: Principal Component Analysis
  36. 36. pc: 2 % variance: 6.073 pc:3%variance:5.441 -0.2 -0.1 0.0 0.1 0.2 -0.2 -0.1 0.0 0.1 0.2 pc: 2 % variance: 6.073 pc:3%variance:5.441 -0.2 -0.1 0.0 0.1 0.2 -0.2 -0.1 0.0 0.1 0.2 Principal Components: PC2 vs PC3 Gp-9 B Gp-9 b
  37. 37. pc: 1 % variance: 12.666 pc:2%variance:6.073 -0.2 -0.1 0.0 0.1 0.2 -0.10 -0.05 0.00 0.05 0.10 0.15 Principal Components: PC1 vs PC2 pc: 1 % variance: 12.666 pc:2%variance:6.073 -0.2 -0.1 0.0 0.1 0.2 -0.10 -0.05 0.00 0.05 0.10 0.15 Gp-9 B ♂ Gp-9 b ♂
  38. 38. brc_m013_0280 brc_m013_0281..brc_m013_0294 brc_m013_0295..brc_m013_0303 brc_m013_0304 brc_m013_0305..brc_m013_0307 brc_m013_0308 brc_m013_0309..brc_m013_0314 brc_m013_0315..brc_m013_0317 brc_m013_0318..brc_m013_0320 brc_m013_0321..brc_m013_0326 brc_m013_0327 brc_m013_0328 brc_m013_0329..brc_m013_0330 brc_m013_0331..brc_m013_0333 brc_m013_0334..brc_m013_0339 brc_m013_0340 brc_m013_0341..brc_m013_0343 brc_m013_0344 brc_m013_0345..brc_m013_0349 brc_m013_0350 brc_m013_0351..brc_m013_0354 brc_m013_0355 brc_m013_0356 brc_m013_0357..brc_m013_0361 brc_m013_0362..brc_m013_0376 brc_m013_0377..brc_m013_0390 brc_m013_0391..brc_m013_0393 brc_m013_0394..brc_m013_0400 brc_m013_0401..brc_m013_0439 brc_m013_0440..brc_m013_0478 brc_m013_0479..brc_m013_0480 140 160 180 brc_m013_0802..brc_m013_0805 brc_m013_0806..brc_m013_0809 brc_m013_0810..brc_m013_0811 brc_m013_0812..brc_m013_0824 brc_m013_0825..brc_m013_0826 brc_m013_0827..brc_m013_0829 brc_m013_0830..brc_m013_0831 brc_m013_0832..brc_m013_0842 brc_m013_0843..brc_m013_0854 brc_m013_0855..brc_m013_0861 brc_m013_0862 brc_m013_0863..brc_m013_0864 brc_m013_0865..brc_m013_0867 brc_m013_0868..brc_m013_0883 brc_m013_0884..brc_m013_0893 brc_m013_0894 brc_m013_0895..brc_m013_0897 brc_m013_0898..brc_m013_0906 brc_m013_0907..brc_m013_0910 brc_m013_0911..brc_m013_0925 brc_m013_0926..brc_m013_0928 brc_m013_0929..brc_m013_0931 140 brc_m013_1275..brc_m013_1280 brc_m013_1281 brc_m013_1282..brc_m013_1286 brc_m013_1287..brc_m013_1298 brc_m013_1299..brc_m013_1307 brc_m013_1308 brc_m013_1309..brc_m013_1313 brc_m013_1314..brc_m013_1317 brc_m013_1318..brc_m013_1319 brc_m013_1320..brc_m013_1326 brc_m013_1327..brc_m013_1340 brc_m013_1341..brc_m013_1362 brc_m013_1363..brc_m013_1385 140 brc_m013_1683 brc_m013_1684 brc_m013_1685..brc_m013_1686 brc_m013_1687..brc_m013_1700 brc_m013_1701..brc_m013_1702 brc_m013_1703 brc_m013_1704..brc_m013_1707 brc_m013_1708..brc_m013_1709 brc_m013_1710..brc_m013_1714 brc_m013_1715..brc_m013_1728 brc_m013_1729..brc_m013_1742 140 brc_m013_1967..brc_m013_1968 brc_m013_1969..brc_m013_1970 brc_m013_1971 brc_m013_1972 brc_m013_1973..brc_m013_1980 brc_m013_1981..brc_m013_1983 brc_m013_1984..brc_m013_1990 brc_m013_1991..brc_m013_1993 brc_m013_1994..brc_m013_1996 brc_m013_1997..brc_m013_2009 brc_m013_2302..brc_m013_2305 brc_m013_2306..brc_m013_2307 brc_m013_2308..brc_m013_2330 brc_m013_2331..brc_m013_2337 brc_m013_2338..brc_m013_2352 brc_m013_2897..brc_m013_2920 brc_m013_2921..brc_m013_2928 brc_m013_2929..brc_m013_2931 brc_m013_2932 brc_m013_2933 brc_m013_2934..brc_m013_2935 brc_m013_2936 brc_m013_2937..brc_m013_2943 brc_m013_2944 brc_m013_2945..brc_m013_2946 brc_m013_2947 brc_m013_2948 brc_m013_2949..brc_m013_2950 brc_m013_2951..brc_m013_2957 brc_m013_2958..brc_m013_2961 brc_m013_2962..brc_m013_2970 brc_m013_2971..brc_m013_2980 brc_m013_2981..brc_m013_2992 brc_m013_2993..brc_m013_2996 brc_m013_2997..brc_m013_2998 brc_m013_2999..brc_m013_3000 brc_m013_3001 brc_m013_3002..brc_m013_3003 brc_m013_3004 brc_m013_3005 brc_m013_3006..brc_m013_3010 brc_m013_3011..brc_m013_3014 brc_m013_3015 brc_m013_3016..brc_m013_3019 brc_m013_3020 brc_m013_3021..brc_m013_3030 brc_m013_3031..brc_m013_3032 brc_m013_3033..brc_m013_3034 brc_m013_3035..brc_m013_3036 brc_m013_3037..brc_m013_3045 brc_m013_3046..brc_m013_3052 brc_m013_3053 brc_m013_3054..brc_m013_3061 brc_m013_3062..brc_m013_3066 brc_m013_3067..brc_m013_3068 brc_m013_3069..brc_m013_3076 brc_m013_3077..brc_m013_3084 brc_m013_3085..brc_m013_3087 brc_m013_3088..brc_m013_3089 brc_m013_3090..brc_m013_3096 brc_m013_3097..brc_m013_3100 brc_m013_3101..brc_m013_3104 brc_m013_3105 brc_m013_3106..brc_m013_3112 brc_m013_3113..brc_m013_3122 brc_m013_3123..brc_m013_3124 brc_m013_3125..brc_m013_3127 brc_m013_3128..brc_m013_3145 brc_m013_3146..brc_m013_3159 brc_m013_3160..brc_m013_3172 0 20 40 60 80 100 LG9 brc_m013_3173..brc_m013_3175 brc_m013_3176..brc_m013_3180 brc_m013_3181..brc_m013_3189 brc_m013_3190..brc_m013_3198 brc_m013_3199 brc_m013_3200..brc_m013_3201 brc_m013_3202..brc_m013_3203 brc_m013_3204 brc_m013_3205..brc_m013_3206 brc_m013_3207..brc_m013_3211 brc_m013_3212..brc_m013_3214 brc_m013_3215..brc_m013_3227 brc_m013_3228..brc_m013_3230 brc_m013_3231..brc_m013_3235 brc_m013_3236..brc_m013_3238 brc_m013_3239..brc_m013_3242 brc_m013_3243..brc_m013_3244 brc_m013_3245 brc_m013_3246..brc_m013_3247 brc_m013_3248..brc_m013_3249 brc_m013_3250..brc_m013_3252 brc_m013_3253..brc_m013_3257 brc_m013_3258 brc_m013_3259 brc_m013_3260..brc_m013_3261 brc_m013_3262..brc_m013_3263 brc_m013_3264 brc_m013_3265..brc_m013_3269 brc_m013_3270..brc_m013_3274 brc_m013_3275..brc_m013_3276 brc_m013_3277..brc_m013_3281 brc_m013_3282..brc_m013_3284 brc_m013_3285 brc_m013_3286..brc_m013_3289 brc_m013_3290..brc_m013_3296 brc_m013_3297 brc_m013_3298..brc_m013_3300 brc_m013_3301..brc_m013_3302 brc_m013_3303..brc_m013_3305 brc_m013_3306..brc_m013_3308 brc_m013_3309..brc_m013_3314 brc_m013_3315..brc_m013_3317 brc_m013_3318..brc_m013_3329 brc_m013_3330..brc_m013_3331 brc_m013_3332..brc_m013_3338 brc_m013_3339..brc_m013_3340 brc_m013_3341..brc_m013_3344 brc_m013_3345..brc_m013_3349 brc_m013_3350..brc_m013_3357 brc_m013_3358..brc_m013_3359 brc_m013_3360 brc_m013_3361..brc_m013_3368 brc_m013_3369..brc_m013_3372 brc_m013_3373..brc_m013_3376 brc_m013_3377 brc_m013_3378..brc_m013_3386 brc_m013_3387..brc_m013_3388 brc_m013_3389..brc_m013_3395 brc_m013_3396..brc_m013_3399 0 20 40 60 80 LG10 brc_m013_3400..brc_m013_3411 brc_m013_3412 brc_m013_3413..brc_m013_3424 brc_m013_3425 brc_m013_3426 brc_m013_3427..brc_m013_3429 brc_m013_3430 brc_m013_3431..brc_m013_3432 brc_m013_3433..brc_m013_3435 brc_m013_3436 brc_m013_3437..brc_m013_3439 brc_m013_3440..brc_m013_3441 brc_m013_3442 brc_m013_3443..brc_m013_3445 brc_m013_3446..brc_m013_3447 brc_m013_3448..brc_m013_3449 brc_m013_3450..brc_m013_3454 brc_m013_3455 brc_m013_3456..brc_m013_3462 brc_m013_3463..brc_m013_3464 brc_m013_3465 brc_m013_3466..brc_m013_3467 brc_m013_3468..brc_m013_3472 brc_m013_3473 brc_m013_3474..brc_m013_3476 brc_m013_3477..brc_m013_3487 brc_m013_3488 brc_m013_3489..brc_m013_3491 brc_m013_3492..brc_m013_3500 brc_m013_3501..brc_m013_3512 brc_m013_3513..brc_m013_3514 brc_m013_3515..brc_m013_3524 brc_m013_3525..brc_m013_3527 brc_m013_3528..brc_m013_3531 brc_m013_3532..brc_m013_3547 brc_m013_3548..brc_m013_3557 brc_m013_3558..brc_m013_3566 brc_m013_3567..brc_m013_3568 brc_m013_3569..brc_m013_3570 brc_m013_3571..brc_m013_3574 brc_m013_3575..brc_m013_3582 brc_m013_3583..brc_m013_3592 brc_m013_3593..brc_m013_3605 brc_m013_3606..brc_m013_3616 brc_m013_3617..brc_m013_3618 brc_m013_3619..brc_m013_3622 brc_m013_3623..brc_m013_3624 brc_m013_3625..brc_m013_3628 brc_m013_3629..brc_m013_3635 0 20 40 60 80 LG11 brc_m013_3636..brc_m013_3661 brc_m013_3662..brc_m013_3665 brc_m013_3666..brc_m013_3667 brc_m013_3668 brc_m013_3669..brc_m013_3671 brc_m013_3672 brc_m013_3673..brc_m013_3674 brc_m013_3675..brc_m013_3682 brc_m013_3683..brc_m013_3685 brc_m013_3686..brc_m013_3688 brc_m013_3689..brc_m013_3693 brc_m013_3694..brc_m013_3698 brc_m013_3699 brc_m013_3700 brc_m013_3701..brc_m013_3702 brc_m013_3703..brc_m013_3704 brc_m013_3705..brc_m013_3712 brc_m013_3713 brc_m013_3714..brc_m013_3716 brc_m013_3717..brc_m013_3724 brc_m013_3725..brc_m013_3730 brc_m013_3731..brc_m013_3752 brc_m013_3753..brc_m013_3758 brc_m013_3759..brc_m013_3789 brc_m013_3790..brc_m013_3801 brc_m013_3802..brc_m013_3814 brc_m013_3815..brc_m013_3818 brc_m013_3819..brc_m013_3822 brc_m013_3823..brc_m013_3826 brc_m013_3827..brc_m013_3832 brc_m013_3833..brc_m013_3837 brc_m013_3838 brc_m013_3839..brc_m013_3841 brc_m013_3842..brc_m013_3847 brc_m013_3848..brc_m013_3853 brc_m013_3854..brc_m013_3858 brc_m013_3859..brc_m013_3868 brc_m013_3869..brc_m013_3871 brc_m013_3872..brc_m013_3901 brc_m013_3902 brc_m013_3903..brc_m013_3909 brc_m013_3910 brc_m013_3911..brc_m013_3926 brc_m013_3927..brc_m013_3931 brc_m013_3932..brc_m013_3948 0 20 40 60 80 LG12 brc_m013_3949..brc_m013_3952 brc_m013_3953..brc_m013_3958 brc_m013_3959..brc_m013_3970 brc_m013_3971 brc_m013_3972..brc_m013_3975 brc_m013_3976 brc_m013_3977..brc_m013_3985 brc_m013_3986 brc_m013_3987..brc_m013_3994 brc_m013_3995..brc_m013_3997 brc_m013_3998..brc_m013_4004 brc_m013_4005..brc_m013_4006 brc_m013_4007..brc_m013_4008 brc_m013_4009..brc_m013_4010 brc_m013_4011..brc_m013_4013 brc_m013_4014 brc_m013_4015 brc_m013_4016..brc_m013_4019 brc_m013_4020..brc_m013_4021 brc_m013_4022..brc_m013_4025 brc_m013_4026..brc_m013_4032 brc_m013_4033..brc_m013_4036 brc_m013_4037..brc_m013_4041 brc_m013_4042..brc_m013_4043 brc_m013_4044..brc_m013_4046 brc_m013_4047..brc_m013_4056 brc_m013_4057 brc_m013_4058..brc_m013_4063 brc_m013_4064..brc_m013_4071 brc_m013_4072..brc_m013_4075 brc_m013_4076 brc_m013_4077 brc_m013_4078..brc_m013_4085 brc_m013_4086..brc_m013_4089 brc_m013_4090..brc_m013_4091 brc_m013_4092..brc_m013_4093 brc_m013_4094..brc_m013_4095 brc_m013_4096..brc_m013_4114 brc_m013_4115..brc_m013_4117 brc_m013_4118..brc_m013_4131 brc_m013_4132..brc_m013_4133 brc_m013_4134..brc_m013_4146 0 20 40 60 80 LG13 brc_m013_4147..brc_m013_4150 brc_m013_4151..brc_m013_4167 brc_m013_4168 brc_m013_4169 brc_m013_4170 brc_m013_4171..brc_m013_4172 brc_m013_4173..brc_m013_4175 brc_m013_4176 brc_m013_4177..brc_m013_4178 brc_m013_4179..brc_m013_4183 brc_m013_4184..brc_m013_4185 brc_m013_4186..brc_m013_4187 brc_m013_4188..brc_m013_4191 brc_m013_4192 brc_m013_4193..brc_m013_4194 brc_m013_4195..brc_m013_4206 brc_m013_4207..brc_m013_4210 brc_m013_4211..brc_m013_4213 brc_m013_4214 brc_m013_4215..brc_m013_4217 brc_m013_4218..brc_m013_4221 brc_m013_4222..brc_m013_4223 brc_m013_4224..brc_m013_4231 brc_m013_4232..brc_m013_4234 brc_m013_4235..brc_m013_4239 brc_m013_4240..brc_m013_4246 brc_m013_4247..brc_m013_4248 brc_m013_4249..brc_m013_4258 brc_m013_4259..brc_m013_4260 brc_m013_4261..brc_m013_4269 brc_m013_4270..brc_m013_4271 brc_m013_4272..brc_m013_4278 brc_m013_4279..brc_m013_4280 brc_m013_4281..brc_m013_4284 brc_m013_4285 brc_m013_4286..brc_m013_4288 brc_m013_4289..brc_m013_4294 brc_m013_4295..brc_m013_4296 brc_m013_4297..brc_m013_4301 brc_m013_4302..brc_m013_4313 brc_m013_4314..brc_m013_4320 brc_m013_4321..brc_m013_4322 brc_m013_4323..brc_m013_4345 brc_m013_4346..brc_m013_4351 0 20 40 60 LG14 brc_m013_4352..brc_m013_4366 brc_m013_4367 brc_m013_4368 brc_m013_4369..brc_m013_4373 brc_m013_4374..brc_m013_4381 brc_m013_4382..brc_m013_4383 brc_m013_4384..brc_m013_4385 brc_m013_4386 brc_m013_4387..brc_m013_4388 brc_m013_4389 brc_m013_4390..brc_m013_4404 brc_m013_4405 brc_m013_4406..brc_m013_4409 brc_m013_4410..brc_m013_4411 brc_m013_4412..brc_m013_4418 brc_m013_4419..brc_m013_4434 brc_m013_4435..brc_m013_4442 brc_m013_4443..brc_m013_4448 brc_m013_4449..brc_m013_4451 brc_m013_4452..brc_m013_4461 brc_m013_4462..brc_m013_4471 brc_m013_4472..brc_m013_4475 brc_m013_4476..brc_m013_4477 brc_m013_4478 brc_m013_4479 brc_m013_4480..brc_m013_4485 brc_m013_4486 brc_m013_4487..brc_m013_4491 brc_m013_4492 brc_m013_4493 brc_m013_4494..brc_m013_4495 brc_m013_4496..brc_m013_4501 brc_m013_4502..brc_m013_4510 brc_m013_4511..brc_m013_4531 brc_m013_4532 brc_m013_4533..brc_m013_4534 brc_m013_4535..brc_m013_4541 brc_m013_4542..brc_m013_4543 brc_m013_4544..brc_m013_4545 brc_m013_4546..brc_m013_4548 brc_m013_4549..brc_m013_4551 brc_m013_4552..brc_m013_4555 brc_m013_4556..brc_m013_4561 0 20 40 60 LG15 brc_m013_4562..brc_m013_4577 brc_m013_4578..brc_m013_4594 brc_m013_4595..brc_m013_4599 brc_m013_4600..brc_m013_4625 brc_m013_4626..brc_m013_4638 brc_m013_4639..brc_m013_4642 brc_m013_4643..brc_m013_4644 brc_m013_4645..brc_m013_4650 brc_m013_4651..brc_m013_4663 brc_m013_4664..brc_m013_4668 brc_m013_4669..brc_m013_4670 brc_m013_4671..brc_m013_4674 brc_m013_4675..brc_m013_4679 brc_m013_4680..brc_m013_4681 brc_m013_4682 brc_m013_4683..brc_m013_4688 brc_m013_4689..brc_m013_4692 brc_m013_4693..brc_m013_4695 brc_m013_4696..brc_m013_4701 brc_m013_4702 brc_m013_4703..brc_m013_4712 brc_m013_4713..brc_m013_4717 brc_m013_4718..brc_m013_4720 brc_m013_4721 brc_m013_4722..brc_m013_4726 brc_m013_4727..brc_m013_4728 brc_m013_4729..brc_m013_4742 brc_m013_4743 brc_m013_4744..brc_m013_4746 brc_m013_4747 brc_m013_4748..brc_m013_4749 brc_m013_4750..brc_m013_4752 brc_m013_4753..brc_m013_4756 brc_m013_4757..brc_m013_4759 brc_m013_4760..brc_m013_4762 brc_m013_4763 brc_m013_4764..brc_m013_4766 brc_m013_4767..brc_m013_4769 brc_m013_4770 brc_m013_4771..brc_m013_4774 brc_m013_4775..brc_m013_4776 brc_m013_4777..brc_m013_4778 brc_m013_4779..brc_m013_4780 brc_m013_4781..brc_m013_4793 brc_m013_4794 brc_m013_4795..brc_m013_4798 brc_m013_4799..brc_m013_4802 brc_m013_4803..brc_m013_4806 brc_m013_4807 brc_m013_4808..brc_m013_4814 brc_m013_4815..brc_m013_4819 brc_m013_4820 brc_m013_4821 brc_m013_4822..brc_m013_4823 brc_m013_4824 brc_m013_4825..brc_m013_4855 brc_m013_4856..brc_m013_4858 brc_m013_4859..brc_m013_4863 brc_m013_4864..brc_m013_4865 brc_m013_4866..brc_m013_4875 brc_m013_4876..brc_m013_4881 brc_m013_4882..brc_m013_4891 brc_m013_4892..brc_m013_4895 brc_m013_4896..brc_m013_4911 brc_m013_4912..brc_m013_4938 brc_m013_4939 brc_m013_4940..brc_m013_4957 brc_m013_4958..brc_m013_4959 brc_m013_4960..brc_m013_4972 brc_m013_4973..brc_m013_4981 brc_m013_4982..brc_m013_4983 0 20 40 60 80 100 120 LGSB Gp-9 Gp-9 brc_m013_0001..brc_m013_0005 brc_m013_0006..brc_m013_0014 brc_m013_0015..brc_m013_0017 brc_m013_0018 brc_m013_0019..brc_m013_0020 brc_m013_0021..brc_m013_0029 brc_m013_0030..brc_m013_0031 brc_m013_0032..brc_m013_0034 brc_m013_0035..brc_m013_0036 brc_m013_0037..brc_m013_0038 brc_m013_0039..brc_m013_0043 brc_m013_0044 brc_m013_0045 brc_m013_0046..brc_m013_0048 brc_m013_0049 brc_m013_0050..brc_m013_0051 brc_m013_0052..brc_m013_0056 brc_m013_0057..brc_m013_0061 brc_m013_0062..brc_m013_0075 brc_m013_0076..brc_m013_0078 brc_m013_0079..brc_m013_0081 brc_m013_0082..brc_m013_0088 brc_m013_0089..brc_m013_0092 brc_m013_0093..brc_m013_0096 brc_m013_0097 brc_m013_0098..brc_m013_0113 brc_m013_0114..brc_m013_0119 brc_m013_0120..brc_m013_0130 brc_m013_0131 brc_m013_0132..brc_m013_0134 brc_m013_0135..brc_m013_0136 brc_m013_0137..brc_m013_0139 brc_m013_0140..brc_m013_0142 brc_m013_0143 brc_m013_0144..brc_m013_0146 brc_m013_0147..brc_m013_0154 brc_m013_0155 brc_m013_0156 brc_m013_0157..brc_m013_0180 brc_m013_0181 brc_m013_0182 brc_m013_0183..brc_m013_0188 brc_m013_0189..brc_m013_0208 brc_m013_0209 brc_m013_0210..brc_m013_0211 brc_m013_0212..brc_m013_0215 brc_m013_0216 brc_m013_0217 brc_m013_0218..brc_m013_0224 brc_m013_0225..brc_m013_0228 brc_m013_0229..brc_m013_0237 brc_m013_0238..brc_m013_0245 brc_m013_0246..brc_m013_0270 brc_m013_0271..brc_m013_0274 brc_m013_0275 brc_m013_0276..brc_m013_0278 brc_m013_0279 brc_m013_0280 brc_m013_0281..brc_m013_0294 brc_m013_0295..brc_m013_0303 brc_m013_0304 brc_m013_0305..brc_m013_0307 brc_m013_0308 brc_m013_0309..brc_m013_0314 brc_m013_0315..brc_m013_0317 brc_m013_0318..brc_m013_0320 brc_m013_0321..brc_m013_0326 brc_m013_0327 brc_m013_0328 brc_m013_0329..brc_m013_0330 brc_m013_0331..brc_m013_0333 brc_m013_0334..brc_m013_0339 brc_m013_0340 brc_m013_0341..brc_m013_0343 brc_m013_0344 brc_m013_0345..brc_m013_0349 brc_m013_0350 brc_m013_0351..brc_m013_0354 brc_m013_0355 brc_m013_0356 brc_m013_0357..brc_m013_0361 brc_m013_0362..brc_m013_0376 brc_m013_0377..brc_m013_0390 brc_m013_0391..brc_m013_0393 brc_m013_0394..brc_m013_0400 brc_m013_0401..brc_m013_0439 brc_m013_0440..brc_m013_0478 brc_m013_0479..brc_m013_0480 0 20 40 60 80 100 120 140 160 180 LG1 brc_m013_0481 brc_m013_0482..brc_m013_0484 brc_m013_0485..brc_m013_0488 brc_m013_0489..brc_m013_0502 brc_m013_0503 brc_m013_0504..brc_m013_0519 brc_m013_0520..brc_m013_0532 brc_m013_0533..brc_m013_0534 brc_m013_0535..brc_m013_0537 brc_m013_0538..brc_m013_0540 brc_m013_0541..brc_m013_0543 brc_m013_0544..brc_m013_0545 brc_m013_0546..brc_m013_0549 brc_m013_0550..brc_m013_0554 brc_m013_0555..brc_m013_0560 brc_m013_0561..brc_m013_0562 brc_m013_0563..brc_m013_0565 brc_m013_0566 brc_m013_0567 brc_m013_0568..brc_m013_0570 brc_m013_0571..brc_m013_0573 brc_m013_0574..brc_m013_0575 brc_m013_0576..brc_m013_0578 brc_m013_0579..brc_m013_0580 brc_m013_0581 brc_m013_0582..brc_m013_0584 brc_m013_0585 brc_m013_0586 brc_m013_0587 brc_m013_0588..brc_m013_0591 brc_m013_0592 brc_m013_0593 brc_m013_0594..brc_m013_0612 brc_m013_0613..brc_m013_0614 brc_m013_0615..brc_m013_0632 brc_m013_0633 brc_m013_0634..brc_m013_0648 brc_m013_0649..brc_m013_0655 brc_m013_0656 brc_m013_0657..brc_m013_0694 brc_m013_0695..brc_m013_0703 brc_m013_0704 brc_m013_0705 brc_m013_0706..brc_m013_0707 brc_m013_0708 brc_m013_0709..brc_m013_0711 brc_m013_0712..brc_m013_0715 brc_m013_0716..brc_m013_0721 brc_m013_0722 brc_m013_0723 brc_m013_0724..brc_m013_0728 brc_m013_0729 brc_m013_0730 brc_m013_0731 brc_m013_0732..brc_m013_0735 brc_m013_0736..brc_m013_0769 brc_m013_0770..brc_m013_0771 brc_m013_0772..brc_m013_0773 brc_m013_0774..brc_m013_0775 brc_m013_0776..brc_m013_0782 brc_m013_0783 brc_m013_0784..brc_m013_0795 brc_m013_0796..brc_m013_0798 brc_m013_0799..brc_m013_0801 brc_m013_0802..brc_m013_0805 brc_m013_0806..brc_m013_0809 brc_m013_0810..brc_m013_0811 brc_m013_0812..brc_m013_0824 brc_m013_0825..brc_m013_0826 brc_m013_0827..brc_m013_0829 brc_m013_0830..brc_m013_0831 brc_m013_0832..brc_m013_0842 brc_m013_0843..brc_m013_0854 brc_m013_0855..brc_m013_0861 brc_m013_0862 brc_m013_0863..brc_m013_0864 brc_m013_0865..brc_m013_0867 brc_m013_0868..brc_m013_0883 brc_m013_0884..brc_m013_0893 brc_m013_0894 brc_m013_0895..brc_m013_0897 brc_m013_0898..brc_m013_0906 brc_m013_0907..brc_m013_0910 brc_m013_0911..brc_m013_0925 brc_m013_0926..brc_m013_0928 brc_m013_0929..brc_m013_0931 0 20 40 60 80 100 120 140 LG2 brc_m013_0932..brc_m013_0941 brc_m013_0942..brc_m013_0943 brc_m013_0944..brc_m013_0945 brc_m013_0946..brc_m013_0949 brc_m013_0950..brc_m013_0952 brc_m013_0953..brc_m013_0975 brc_m013_0976..brc_m013_1019 brc_m013_1020 brc_m013_1021 brc_m013_1022..brc_m013_1061 brc_m013_1062 brc_m013_1063 brc_m013_1064..brc_m013_1065 brc_m013_1066..brc_m013_1068 brc_m013_1069 brc_m013_1070 brc_m013_1071..brc_m013_1074 brc_m013_1075 brc_m013_1076..brc_m013_1081 brc_m013_1082..brc_m013_1086 brc_m013_1087..brc_m013_1088 brc_m013_1089..brc_m013_1098 brc_m013_1099..brc_m013_1106 brc_m013_1107..brc_m013_1116 brc_m013_1117 brc_m013_1118..brc_m013_1121 brc_m013_1122..brc_m013_1127 brc_m013_1128 brc_m013_1129..brc_m013_1136 brc_m013_1137..brc_m013_1138 brc_m013_1139..brc_m013_1141 brc_m013_1142..brc_m013_1144 brc_m013_1145..brc_m013_1156 brc_m013_1157 brc_m013_1158..brc_m013_1170 brc_m013_1171..brc_m013_1181 brc_m013_1182..brc_m013_1185 brc_m013_1186 brc_m013_1187..brc_m013_1205 brc_m013_1206..brc_m013_1218 brc_m013_1219..brc_m013_1220 brc_m013_1221..brc_m013_1224 brc_m013_1225..brc_m013_1228 brc_m013_1229 brc_m013_1230..brc_m013_1236 brc_m013_1237 brc_m013_1238..brc_m013_1247 brc_m013_1248..brc_m013_1251 brc_m013_1252 brc_m013_1253..brc_m013_1268 brc_m013_1269..brc_m013_1270 brc_m013_1271..brc_m013_1273 brc_m013_1274 brc_m013_1275..brc_m013_1280 brc_m013_1281 brc_m013_1282..brc_m013_1286 brc_m013_1287..brc_m013_1298 brc_m013_1299..brc_m013_1307 brc_m013_1308 brc_m013_1309..brc_m013_1313 brc_m013_1314..brc_m013_1317 brc_m013_1318..brc_m013_1319 brc_m013_1320..brc_m013_1326 brc_m013_1327..brc_m013_1340 brc_m013_1341..brc_m013_1362 brc_m013_1363..brc_m013_1385 0 20 40 60 80 100 120 140 LG3 brc_m013_1386..brc_m013_1388 brc_m013_1389..brc_m013_1398 brc_m013_1399..brc_m013_1406 brc_m013_1407..brc_m013_1411 brc_m013_1412..brc_m013_1413 brc_m013_1414..brc_m013_1416 brc_m013_1417 brc_m013_1418..brc_m013_1420 brc_m013_1421..brc_m013_1424 brc_m013_1425..brc_m013_1432 brc_m013_1433..brc_m013_1442 brc_m013_1443 brc_m013_1444..brc_m013_1450 brc_m013_1451 brc_m013_1452 brc_m013_1453..brc_m013_1455 brc_m013_1456..brc_m013_1467 brc_m013_1468..brc_m013_1469 brc_m013_1470 brc_m013_1471..brc_m013_1474 brc_m013_1475 brc_m013_1476 brc_m013_1477 brc_m013_1478..brc_m013_1482 brc_m013_1483 brc_m013_1484 brc_m013_1485..brc_m013_1487 brc_m013_1488..brc_m013_1490 brc_m013_1491 brc_m013_1492..brc_m013_1494 brc_m013_1495..brc_m013_1496 brc_m013_1497..brc_m013_1500 brc_m013_1501 brc_m013_1502..brc_m013_1513 brc_m013_1514..brc_m013_1562 brc_m013_1563..brc_m013_1565 brc_m013_1566..brc_m013_1567 brc_m013_1568..brc_m013_1580 brc_m013_1581..brc_m013_1587 brc_m013_1588..brc_m013_1591 brc_m013_1592..brc_m013_1593 brc_m013_1594..brc_m013_1604 brc_m013_1605..brc_m013_1607 brc_m013_1608..brc_m013_1609 brc_m013_1610..brc_m013_1611 brc_m013_1612..brc_m013_1616 brc_m013_1617..brc_m013_1618 brc_m013_1619..brc_m013_1620 brc_m013_1621..brc_m013_1629 brc_m013_1630..brc_m013_1633 brc_m013_1634..brc_m013_1638 brc_m013_1639..brc_m013_1647 brc_m013_1648..brc_m013_1649 brc_m013_1650..brc_m013_1656 brc_m013_1657..brc_m013_1665 brc_m013_1666..brc_m013_1672 brc_m013_1673..brc_m013_1674 brc_m013_1675..brc_m013_1678 brc_m013_1679..brc_m013_1682 brc_m013_1683 brc_m013_1684 brc_m013_1685..brc_m013_1686 brc_m013_1687..brc_m013_1700 brc_m013_1701..brc_m013_1702 brc_m013_1703 brc_m013_1704..brc_m013_1707 brc_m013_1708..brc_m013_1709 brc_m013_1710..brc_m013_1714 brc_m013_1715..brc_m013_1728 brc_m013_1729..brc_m013_1742 0 20 40 60 80 100 120 140 LG4 brc_m013_1743..brc_m013_1750 brc_m013_1751..brc_m013_1766 brc_m013_1767 brc_m013_1768 brc_m013_1769..brc_m013_1772 brc_m013_1773..brc_m013_1779 brc_m013_1780 brc_m013_1781 brc_m013_1782..brc_m013_1783 brc_m013_1784 brc_m013_1785..brc_m013_1786 brc_m013_1787 brc_m013_1788 brc_m013_1789..brc_m013_1790 brc_m013_1791..brc_m013_1793 brc_m013_1794..brc_m013_1797 brc_m013_1798..brc_m013_1800 brc_m013_1801..brc_m013_1804 brc_m013_1805 brc_m013_1806..brc_m013_1808 brc_m013_1809 brc_m013_1810..brc_m013_1813 brc_m013_1814..brc_m013_1818 brc_m013_1819..brc_m013_1820 brc_m013_1821..brc_m013_1822 brc_m013_1823..brc_m013_1824 brc_m013_1825 brc_m013_1826..brc_m013_1840 brc_m013_1841..brc_m013_1842 brc_m013_1843 brc_m013_1844..brc_m013_1848 brc_m013_1849 brc_m013_1850 brc_m013_1851..brc_m013_1853 brc_m013_1854..brc_m013_1858 brc_m013_1859..brc_m013_1866 brc_m013_1867..brc_m013_1868 brc_m013_1869 brc_m013_1870..brc_m013_1874 brc_m013_1875..brc_m013_1876 brc_m013_1877..brc_m013_1878 brc_m013_1879..brc_m013_1883 brc_m013_1884 brc_m013_1885 brc_m013_1886..brc_m013_1888 brc_m013_1889..brc_m013_1895 brc_m013_1896..brc_m013_1899 brc_m013_1900..brc_m013_1913 brc_m013_1914 brc_m013_1915..brc_m013_1922 brc_m013_1923 brc_m013_1924..brc_m013_1928 brc_m013_1929..brc_m013_1942 brc_m013_1943..brc_m013_1946 brc_m013_1947..brc_m013_1948 brc_m013_1949 brc_m013_1950..brc_m013_1953 brc_m013_1954..brc_m013_1955 brc_m013_1956 brc_m013_1957..brc_m013_1963 brc_m013_1964..brc_m013_1966 brc_m013_1967..brc_m013_1968 brc_m013_1969..brc_m013_1970 brc_m013_1971 brc_m013_1972 brc_m013_1973..brc_m013_1980 brc_m013_1981..brc_m013_1983 brc_m013_1984..brc_m013_1990 brc_m013_1991..brc_m013_1993 brc_m013_1994..brc_m013_1996 brc_m013_1997..brc_m013_2009 0 20 40 60 80 100 120 LG5 brc_m013_2010..brc_m013_2028 brc_m013_2029..brc_m013_2038 brc_m013_2039 brc_m013_2040..brc_m013_2041 brc_m013_2042..brc_m013_2047 brc_m013_2048..brc_m013_2050 brc_m013_2051..brc_m013_2053 brc_m013_2054..brc_m013_2062 brc_m013_2063 brc_m013_2064..brc_m013_2065 brc_m013_2066..brc_m013_2067 brc_m013_2068 brc_m013_2069..brc_m013_2071 brc_m013_2072..brc_m013_2081 brc_m013_2082 brc_m013_2083 brc_m013_2084 brc_m013_2085..brc_m013_2099 brc_m013_2100 brc_m013_2101..brc_m013_2102 brc_m013_2103..brc_m013_2107 brc_m013_2108 brc_m013_2109..brc_m013_2112 brc_m013_2113..brc_m013_2114 brc_m013_2115..brc_m013_2123 brc_m013_2124..brc_m013_2131 brc_m013_2132 brc_m013_2133 brc_m013_2134..brc_m013_2136 brc_m013_2137 brc_m013_2138..brc_m013_2139 brc_m013_2140..brc_m013_2142 brc_m013_2143..brc_m013_2150 brc_m013_2151..brc_m013_2152 brc_m013_2153..brc_m013_2161 brc_m013_2162..brc_m013_2163 brc_m013_2164..brc_m013_2165 brc_m013_2166..brc_m013_2170 brc_m013_2171..brc_m013_2172 brc_m013_2173 brc_m013_2174..brc_m013_2182 brc_m013_2183..brc_m013_2186 brc_m013_2187..brc_m013_2190 brc_m013_2191..brc_m013_2193 brc_m013_2194 brc_m013_2195..brc_m013_2201 brc_m013_2202..brc_m013_2203 brc_m013_2204..brc_m013_2220 brc_m013_2221..brc_m013_2232 brc_m013_2233..brc_m013_2239 brc_m013_2240..brc_m013_2261 brc_m013_2262..brc_m013_2267 brc_m013_2268..brc_m013_2269 brc_m013_2270..brc_m013_2271 brc_m013_2272..brc_m013_2282 brc_m013_2283..brc_m013_2284 brc_m013_2285..brc_m013_2299 brc_m013_2300..brc_m013_2301 brc_m013_2302..brc_m013_2305 brc_m013_2306..brc_m013_2307 brc_m013_2308..brc_m013_2330 brc_m013_2331..brc_m013_2337 brc_m013_2338..brc_m013_2352 0 20 40 60 80 100 120 LG6 brc_m013_2353..brc_m013_2365 brc_m013_2366..brc_m013_2369 brc_m013_2370..brc_m013_2372 brc_m013_2373..brc_m013_2378 brc_m013_2379..brc_m013_2386 brc_m013_2387 brc_m013_2388..brc_m013_2394 brc_m013_2395..brc_m013_2397 brc_m013_2398 brc_m013_2399 brc_m013_2400 brc_m013_2401 brc_m013_2402..brc_m013_2407 brc_m013_2408..brc_m013_2411 brc_m013_2412..brc_m013_2416 brc_m013_2417 brc_m013_2418 brc_m013_2419..brc_m013_2436 brc_m013_2437..brc_m013_2441 brc_m013_2442 brc_m013_2443..brc_m013_2444 brc_m013_2445 brc_m013_2446..brc_m013_2453 brc_m013_2454 brc_m013_2455..brc_m013_2460 brc_m013_2461 brc_m013_2462..brc_m013_2470 brc_m013_2471..brc_m013_2474 brc_m013_2475..brc_m013_2482 brc_m013_2483 brc_m013_2484..brc_m013_2487 brc_m013_2488 brc_m013_2489..brc_m013_2492 brc_m013_2493..brc_m013_2496 brc_m013_2497 brc_m013_2498..brc_m013_2504 brc_m013_2505 brc_m013_2506..brc_m013_2510 brc_m013_2511..brc_m013_2523 brc_m013_2524..brc_m013_2531 brc_m013_2532..brc_m013_2536 brc_m013_2537..brc_m013_2555 brc_m013_2556..brc_m013_2571 brc_m013_2572..brc_m013_2573 brc_m013_2574..brc_m013_2579 brc_m013_2580..brc_m013_2581 brc_m013_2582..brc_m013_2587 brc_m013_2588 brc_m013_2589..brc_m013_2594 brc_m013_2595 brc_m013_2596..brc_m013_2597 brc_m013_2598..brc_m013_2604 brc_m013_2605 brc_m013_2606..brc_m013_2616 brc_m013_2617..brc_m013_2619 brc_m013_2620..brc_m013_2623 brc_m013_2624 brc_m013_2625..brc_m013_2626 brc_m013_2627..brc_m013_2628 brc_m013_2629..brc_m013_2630 0 20 40 60 80 100 LG7 brc_m013_2631..brc_m013_2632 brc_m013_2633 brc_m013_2634..brc_m013_2635 brc_m013_2636..brc_m013_2642 brc_m013_2643..brc_m013_2657 brc_m013_2658..brc_m013_2659 brc_m013_2660..brc_m013_2661 brc_m013_2662 brc_m013_2663 brc_m013_2664 brc_m013_2665..brc_m013_2666 brc_m013_2667..brc_m013_2668 brc_m013_2669..brc_m013_2670 brc_m013_2671..brc_m013_2680 brc_m013_2681..brc_m013_2682 brc_m013_2683 brc_m013_2684..brc_m013_2685 brc_m013_2686..brc_m013_2694 brc_m013_2695..brc_m013_2698 brc_m013_2699..brc_m013_2713 brc_m013_2714..brc_m013_2725 brc_m013_2726..brc_m013_2727 brc_m013_2728..brc_m013_2731 brc_m013_2732 brc_m013_2733..brc_m013_2753 brc_m013_2754..brc_m013_2758 brc_m013_2759..brc_m013_2763 brc_m013_2764..brc_m013_2779 brc_m013_2780 brc_m013_2781 brc_m013_2782..brc_m013_2784 brc_m013_2785..brc_m013_2787 brc_m013_2788..brc_m013_2791 brc_m013_2792..brc_m013_2797 brc_m013_2798..brc_m013_2799 brc_m013_2800 brc_m013_2801..brc_m013_2804 brc_m013_2805..brc_m013_2809 brc_m013_2810..brc_m013_2811 brc_m013_2812..brc_m013_2813 brc_m013_2814..brc_m013_2817 brc_m013_2818..brc_m013_2827 brc_m013_2828 brc_m013_2829..brc_m013_2832 brc_m013_2833 brc_m013_2834..brc_m013_2840 brc_m013_2841..brc_m013_2846 brc_m013_2847 brc_m013_2848..brc_m013_2852 brc_m013_2853 brc_m013_2854..brc_m013_2856 brc_m013_2857..brc_m013_2862 brc_m013_2863..brc_m013_2868 brc_m013_2869..brc_m013_2874 brc_m013_2875..brc_m013_2896 0 20 40 60 80 100 LG8 brc_m013_2897..brc_m013_2920 brc_m013_2921..brc_m013_2928 brc_m013_2929..brc_m013_2931 brc_m013_2932 brc_m013_2933 brc_m013_2934..brc_m013_2935 brc_m013_2936 brc_m013_2937..brc_m013_2943 brc_m013_2944 brc_m013_2945..brc_m013_2946 brc_m013_2947 brc_m013_2948 brc_m013_2949..brc_m013_2950 brc_m013_2951..brc_m013_2957 brc_m013_2958..brc_m013_2961 brc_m013_2962..brc_m013_2970 brc_m013_2971..brc_m013_2980 brc_m013_2981..brc_m013_2992 brc_m013_2993..brc_m013_2996 brc_m013_2997..brc_m013_2998 brc_m013_2999..brc_m013_3000 brc_m013_3001 brc_m013_3002..brc_m013_3003 brc_m013_3004 brc_m013_3005 brc_m013_3006..brc_m013_3010 brc_m013_3011..brc_m013_3014 brc_m013_3015 brc_m013_3016..brc_m013_3019 brc_m013_3020 brc_m013_3021..brc_m013_3030 brc_m013_3031..brc_m013_3032 brc_m013_3033..brc_m013_3034 brc_m013_3035..brc_m013_3036 brc_m013_3037..brc_m013_3045 brc_m013_3046..brc_m013_3052 brc_m013_3053 brc_m013_3054..brc_m013_3061 brc_m013_3062..brc_m013_3066 brc_m013_3067..brc_m013_3068 brc_m013_3069..brc_m013_3076 brc_m013_3077..brc_m013_3084 brc_m013_3085..brc_m013_3087 brc_m013_3088..brc_m013_3089 brc_m013_3090..brc_m013_3096 brc_m013_3097..brc_m013_3100 brc_m013_3101..brc_m013_3104 brc_m013_3105 brc_m013_3106..brc_m013_3112 brc_m013_3113..brc_m013_3122 brc_m013_3123..brc_m013_3124 brc_m013_3125..brc_m013_3127 brc_m013_3128..brc_m013_3145 brc_m013_3146..brc_m013_3159 brc_m013_3160..brc_m013_3172 0 20 40 60 80 100 LG9 brc_m013_3173..brc_m013_3175 brc_m013_3176..brc_m013_3180 brc_m013_3181..brc_m013_3189 brc_m013_3190..brc_m013_3198 brc_m013_3199 brc_m013_3200..brc_m013_3201 brc_m013_3202..brc_m013_3203 brc_m013_3204 brc_m013_3205..brc_m013_3206 brc_m013_3207..brc_m013_3211 brc_m013_3212..brc_m013_3214 brc_m013_3215..brc_m013_3227 brc_m013_3228..brc_m013_3230 brc_m013_3231..brc_m013_3235 brc_m013_3236..brc_m013_3238 brc_m013_3239..brc_m013_3242 brc_m013_3243..brc_m013_3244 brc_m013_3245 brc_m013_3246..brc_m013_3247 brc_m013_3248..brc_m013_3249 brc_m013_3250..brc_m013_3252 brc_m013_3253..brc_m013_3257 brc_m013_3258 brc_m013_3259 brc_m013_3260..brc_m013_3261 brc_m013_3262..brc_m013_3263 brc_m013_3264 brc_m013_3265..brc_m013_3269 brc_m013_3270..brc_m013_3274 brc_m013_3275..brc_m013_3276 brc_m013_3277..brc_m013_3281 brc_m013_3282..brc_m013_3284 brc_m013_3285 brc_m013_3286..brc_m013_3289 brc_m013_3290..brc_m013_3296 brc_m013_3297 brc_m013_3298..brc_m013_3300 brc_m013_3301..brc_m013_3302 brc_m013_3303..brc_m013_3305 brc_m013_3306..brc_m013_3308 brc_m013_3309..brc_m013_3314 brc_m013_3315..brc_m013_3317 brc_m013_3318..brc_m013_3329 brc_m013_3330..brc_m013_3331 brc_m013_3332..brc_m013_3338 brc_m013_3339..brc_m013_3340 brc_m013_3341..brc_m013_3344 brc_m013_3345..brc_m013_3349 brc_m013_3350..brc_m013_3357 brc_m013_3358..brc_m013_3359 brc_m013_3360 brc_m013_3361..brc_m013_3368 brc_m013_3369..brc_m013_3372 brc_m013_3373..brc_m013_3376 brc_m013_3377 brc_m013_3378..brc_m013_3386 brc_m013_3387..brc_m013_3388 brc_m013_3389..brc_m013_3395 brc_m013_3396..brc_m013_3399 0 20 40 60 80 LG10 brc_m013_3400..brc_m013_3411 brc_m013_3412 brc_m013_3413..brc_m013_3424 brc_m013_3425 brc_m013_3426 brc_m013_3427..brc_m013_3429 brc_m013_3430 brc_m013_3431..brc_m013_3432 brc_m013_3433..brc_m013_3435 brc_m013_3436 brc_m013_3437..brc_m013_3439 brc_m013_3440..brc_m013_3441 brc_m013_3442 brc_m013_3443..brc_m013_3445 brc_m013_3446..brc_m013_3447 brc_m013_3448..brc_m013_3449 brc_m013_3450..brc_m013_3454 brc_m013_3455 brc_m013_3456..brc_m013_3462 brc_m013_3463..brc_m013_3464 brc_m013_3465 brc_m013_3466..brc_m013_3467 brc_m013_3468..brc_m013_3472 brc_m013_3473 brc_m013_3474..brc_m013_3476 brc_m013_3477..brc_m013_3487 brc_m013_3488 brc_m013_3489..brc_m013_3491 brc_m013_3492..brc_m013_3500 brc_m013_3501..brc_m013_3512 brc_m013_3513..brc_m013_3514 brc_m013_3515..brc_m013_3524 brc_m013_3525..brc_m013_3527 brc_m013_3528..brc_m013_3531 brc_m013_3532..brc_m013_3547 brc_m013_3548..brc_m013_3557 brc_m013_3558..brc_m013_3566 brc_m013_3567..brc_m013_3568 brc_m013_3569..brc_m013_3570 brc_m013_3571..brc_m013_3574 brc_m013_3575..brc_m013_3582 brc_m013_3583..brc_m013_3592 brc_m013_3593..brc_m013_3605 brc_m013_3606..brc_m013_3616 brc_m013_3617..brc_m013_3618 brc_m013_3619..brc_m013_3622 brc_m013_3623..brc_m013_3624 brc_m013_3625..brc_m013_3628 brc_m013_3629..brc_m013_3635 0 20 40 60 80 LG11 brc_m013_3636..brc_m013_3661 brc_m013_3662..brc_m013_3665 brc_m013_3666..brc_m013_3667 brc_m013_3668 brc_m013_3669..brc_m013_3671 brc_m013_3672 brc_m013_3673..brc_m013_3674 brc_m013_3675..brc_m013_3682 brc_m013_3683..brc_m013_3685 brc_m013_3686..brc_m013_3688 brc_m013_3689..brc_m013_3693 brc_m013_3694..brc_m013_3698 brc_m013_3699 brc_m013_3700 brc_m013_3701..brc_m013_3702 brc_m013_3703..brc_m013_3704 brc_m013_3705..brc_m013_3712 brc_m013_3713 brc_m013_3714..brc_m013_3716 brc_m013_3717..brc_m013_3724 brc_m013_3725..brc_m013_3730 brc_m013_3731..brc_m013_3752 brc_m013_3753..brc_m013_3758 brc_m013_3759..brc_m013_3789 brc_m013_3790..brc_m013_3801 brc_m013_3802..brc_m013_3814 brc_m013_3815..brc_m013_3818 brc_m013_3819..brc_m013_3822 brc_m013_3823..brc_m013_3826 brc_m013_3827..brc_m013_3832 brc_m013_3833..brc_m013_3837 brc_m013_3838 brc_m013_3839..brc_m013_3841 brc_m013_3842..brc_m013_3847 brc_m013_3848..brc_m013_3853 brc_m013_3854..brc_m013_3858 brc_m013_3859..brc_m013_3868 brc_m013_3869..brc_m013_3871 brc_m013_3872..brc_m013_3901 brc_m013_3902 brc_m013_3903..brc_m013_3909 brc_m013_3910 brc_m013_3911..brc_m013_3926 brc_m013_3927..brc_m013_3931 brc_m013_3932..brc_m013_3948 0 20 40 60 80 LG12 brc_m013_3949..brc_m013_3952 brc_m013_3953..brc_m013_3958 brc_m013_3959..brc_m013_3970 brc_m013_3971 brc_m013_3972..brc_m013_3975 brc_m013_3976 brc_m013_3977..brc_m013_3985 brc_m013_3986 brc_m013_3987..brc_m013_3994 brc_m013_3995..brc_m013_3997 brc_m013_3998..brc_m013_4004 brc_m013_4005..brc_m013_4006 brc_m013_4007..brc_m013_4008 brc_m013_4009..brc_m013_4010 brc_m013_4011..brc_m013_4013 brc_m013_4014 brc_m013_4015 brc_m013_4016..brc_m013_4019 brc_m013_4020..brc_m013_4021 brc_m013_4022..brc_m013_4025 brc_m013_4026..brc_m013_4032 brc_m013_4033..brc_m013_4036 brc_m013_4037..brc_m013_4041 brc_m013_4042..brc_m013_4043 brc_m013_4044..brc_m013_4046 brc_m013_4047..brc_m013_4056 brc_m013_4057 brc_m013_4058..brc_m013_4063 brc_m013_4064..brc_m013_4071 brc_m013_4072..brc_m013_4075 brc_m013_4076 brc_m013_4077 brc_m013_4078..brc_m013_4085 brc_m013_4086..brc_m013_4089 brc_m013_4090..brc_m013_4091 brc_m013_4092..brc_m013_4093 brc_m013_4094..brc_m013_4095 brc_m013_4096..brc_m013_4114 brc_m013_4115..brc_m013_4117 brc_m013_4118..brc_m013_4131 brc_m013_4132..brc_m013_4133 brc_m013_4134..brc_m013_4146 0 20 40 60 80 LG13 brc_m013_4147..brc_m013_4150 brc_m013_4151..brc_m013_4167 brc_m013_4168 brc_m013_4169 brc_m013_4170 brc_m013_4171..brc_m013_4172 brc_m013_4173..brc_m013_4175 brc_m013_4176 brc_m013_4177..brc_m013_4178 brc_m013_4179..brc_m013_4183 brc_m013_4184..brc_m013_4185 brc_m013_4186..brc_m013_4187 brc_m013_4188..brc_m013_4191 brc_m013_4192 brc_m013_4193..brc_m013_4194 brc_m013_4195..brc_m013_4206 brc_m013_4207..brc_m013_4210 brc_m013_4211..brc_m013_4213 brc_m013_4214 brc_m013_4215..brc_m013_4217 brc_m013_4218..brc_m013_4221 brc_m013_4222..brc_m013_4223 brc_m013_4224..brc_m013_4231 brc_m013_4232..brc_m013_4234 brc_m013_4235..brc_m013_4239 brc_m013_4240..brc_m013_4246 brc_m013_4247..brc_m013_4248 brc_m013_4249..brc_m013_4258 brc_m013_4259..brc_m013_4260 brc_m013_4261..brc_m013_4269 brc_m013_4270..brc_m013_4271 brc_m013_4272..brc_m013_4278 brc_m013_4279..brc_m013_4280 brc_m013_4281..brc_m013_4284 brc_m013_4285 brc_m013_4286..brc_m013_4288 brc_m013_4289..brc_m013_4294 brc_m013_4295..brc_m013_4296 brc_m013_4297..brc_m013_4301 brc_m013_4302..brc_m013_4313 brc_m013_4314..brc_m013_4320 brc_m013_4321..brc_m013_4322 brc_m013_4323..brc_m013_4345 brc_m013_4346..brc_m013_4351 0 20 40 60 LG14 brc_m013_4352..brc_m013_4366 brc_m013_4367 brc_m013_4368 brc_m013_4369..brc_m013_4373 brc_m013_4374..brc_m013_4381 brc_m013_4382..brc_m013_4383 brc_m013_4384..brc_m013_4385 brc_m013_4386 brc_m013_4387..brc_m013_4388 brc_m013_4389 brc_m013_4390..brc_m013_4404 brc_m013_4405 brc_m013_4406..brc_m013_4409 brc_m013_4410..brc_m013_4411 brc_m013_4412..brc_m013_4418 brc_m013_4419..brc_m013_4434 brc_m013_4435..brc_m013_4442 brc_m013_4443..brc_m013_4448 brc_m013_4449..brc_m013_4451 brc_m013_4452..brc_m013_4461 brc_m013_4462..brc_m013_4471 brc_m013_4472..brc_m013_4475 brc_m013_4476..brc_m013_4477 brc_m013_4478 brc_m013_4479 brc_m013_4480..brc_m013_4485 brc_m013_4486 brc_m013_4487..brc_m013_4491 brc_m013_4492 brc_m013_4493 brc_m013_4494..brc_m013_4495 brc_m013_4496..brc_m013_4501 brc_m013_4502..brc_m013_4510 brc_m013_4511..brc_m013_4531 brc_m013_4532 brc_m013_4533..brc_m013_4534 brc_m013_4535..brc_m013_4541 brc_m013_4542..brc_m013_4543 brc_m013_4544..brc_m013_4545 brc_m013_4546..brc_m013_4548 brc_m013_4549..brc_m013_4551 brc_m013_4552..brc_m013_4555 brc_m013_4556..brc_m013_4561 0 20 40 60 LG15 brc_m013_4562..brc_m013_4577 brc_m013_4578..brc_m013_4594 brc_m013_4595..brc_m013_4599 brc_m013_4600..brc_m013_4625 brc_m013_4626..brc_m013_4638 brc_m013_4639..brc_m013_4642 brc_m013_4643..brc_m013_4644 brc_m013_4645..brc_m013_4650 brc_m013_4651..brc_m013_4663 brc_m013_4664..brc_m013_4668 brc_m013_4669..brc_m013_4670 brc_m013_4671..brc_m013_4674 brc_m013_4675..brc_m013_4679 brc_m013_4680..brc_m013_4681 brc_m013_4682 brc_m013_4683..brc_m013_4688 brc_m013_4689..brc_m013_4692 brc_m013_4693..brc_m013_4695 brc_m013_4696..brc_m013_4701 brc_m013_4702 brc_m013_4703..brc_m013_4712 brc_m013_4713..brc_m013_4717 brc_m013_4718..brc_m013_4720 brc_m013_4721 brc_m013_4722..brc_m013_4726 brc_m013_4727..brc_m013_4728 brc_m013_4729..brc_m013_4742 brc_m013_4743 brc_m013_4744..brc_m013_4746 brc_m013_4747 brc_m013_4748..brc_m013_4749 brc_m013_4750..brc_m013_4752 brc_m013_4753..brc_m013_4756 brc_m013_4757..brc_m013_4759 brc_m013_4760..brc_m013_4762 brc_m013_4763 brc_m013_4764..brc_m013_4766 brc_m013_4767..brc_m013_4769 brc_m013_4770 brc_m013_4771..brc_m013_4774 brc_m013_4775..brc_m013_4776 brc_m013_4777..brc_m013_4778 brc_m013_4779..brc_m013_4780 brc_m013_4781..brc_m013_4793 brc_m013_4794 brc_m013_4795..brc_m013_4798 brc_m013_4799..brc_m013_4802 brc_m013_4803..brc_m013_4806 brc_m013_4807 brc_m013_4808..brc_m013_4814 brc_m013_4815..brc_m013_4819 brc_m013_4820 brc_m013_4821 brc_m013_4822..brc_m013_4823 brc_m013_4824 brc_m013_4825..brc_m013_4855 brc_m013_4856..brc_m013_4858 brc_m013_4859..brc_m013_4863 brc_m013_4864..brc_m013_4865 brc_m013_4866..brc_m013_4875 brc_m013_4876..brc_m013_4881 brc_m013_4882..brc_m013_4891 brc_m013_4892..brc_m013_4895 brc_m013_4896..brc_m013_4911 brc_m013_4912..brc_m013_4938 brc_m013_4939 brc_m013_4940..brc_m013_4957 brc_m013_4958..brc_m013_4959 brc_m013_4960..brc_m013_4972 brc_m013_4973..brc_m013_4981 brc_m013_4982..brc_m013_4983 0 20 40 60 80 100 120 LGSB Gp-9 Figure 1 a b Si_gnF.scaffold00779_nt2778431.7 Si_gnF.scaffold00779_nt1255229 Si_gnF.scaffold02684_nt10884.2 Si_gnF.scaffold00779_nt1633919 Si_gnF.scaffold00779_nt17842486.5 Si_gnF.scaffold00779_nt3746833 Si_gnF.scaffold00779_nt374687927.3 Si_gnF.scaffold00779_nt382158728.5 Si_gnF.scaffold00779_nt417489034.2 Si_gnF.scaffold09607_nt698300 Si_gnF.scaffold09607_nt69848340.2 Si_gnF.scaffold09758_nt22273252.8 Si_gnF.scaffold05266_nt63430678.4 Si_gnF.scaffold05266_nt65952779.7 Si_gnF.scaffold05266_nt733643 Si_gnF.scaffold05266_nt75364482.8 Si_gnF.scaffold07090_nt71001087.6 Si_gnF.scaffold07090_nt105177192.7 Si_gnF.scaffold00255_nt314067 Si_gnF.scaffold00255_nt40777897.5 Si_gnF.scaffold03404_nt128925 Si_gnF.scaffold03404_nt228606 Si_gnF.scaffold03404_nt241461 104.4 Si_gnF.scaffold00413_nt676115107.3 Si_gnF.scaffold00413_nt1035856109.6 Si_gnF.scaffold01573_nt108462110.8 Si_gnF.scaffold01573_nt447618112.1 Si_gnF.scaffold00899_nt377419 Si_gnF.scaffold00899_nt686574114.4 Si_gnF.scaffold00899_nt236146 Si_gnF.scaffold00899_nt332715 Si_gnF.scaffold00899_nt335756 115.8 Si_gnF.scaffold00469_nt794118.0 Si_gnF.scaffold00690_nt229012 Si_gnF.scaffold00690_nt415290119.3 Si_gnF.scaffold06914_nt297673125.1 Si_gnF.scaffold01957_nt412242 Si_gnF.scaffold02848_nt41846127.6 LGS B from M013 Si_gnF.scaffold00779_nt2778430.0 Si_gnF.scaffold00779_nt1255229 Si_gnF.scaffold02684_nt10881.2 Si_gnF.scaffold00779_nt1633919 Si_gnF.scaffold00779_nt17842486.7 Si_gnF.scaffold00779_nt3746833 Si_gnF.scaffold00779_nt3746879 Si_gnF.scaffold00779_nt3821587 22.2 Si_gnF.scaffold00779_nt417489029.2 Si_gnF.scaffold09607_nt698300 Si_gnF.scaffold09607_nt69848361.9 Si_gnF.scaffold01573_nt10846280.7 Si_gnF.scaffold00255_nt314067 Si_gnF.scaffold00255_nt407778 Si_gnF.scaffold00413_nt1035856 Si_gnF.scaffold00413_nt676115 Si_gnF.scaffold00469_nt794 Si_gnF.scaffold00690_nt229012 Si_gnF.scaffold00690_nt415290 Si_gnF.scaffold00899_nt236146 Si_gnF.scaffold00899_nt332715 Si_gnF.scaffold00899_nt335756 Si_gnF.scaffold00899_nt377419 Si_gnF.scaffold00899_nt686574 Si_gnF.scaffold01573_nt447618 Si_gnF.scaffold01957_nt412242 Si_gnF.scaffold03404_nt128925 Si_gnF.scaffold03404_nt228606 Si_gnF.scaffold03404_nt241461 Si_gnF.scaffold05266_nt634306 Si_gnF.scaffold05266_nt659527 Si_gnF.scaffold05266_nt733643 Si_gnF.scaffold05266_nt753644 Si_gnF.scaffold06914_nt297673 Si_gnF.scaffold07090_nt1051771 Si_gnF.scaffold07090_nt710010 Si_gnF.scaffold09758_nt222732 81.8 Si_gnF.scaffold02848_nt4184690.5 LGS B/b from P034 Gp-9 Total 285 non- recombining markers >4% of genome linked to Gp-9 No recombination between B and b over ⅔ of a chromosme! Gp-9 Wang & Wurm et al 2013 Nature
  39. 39. • Is this gene the single überregulator? maybe 1/14th of the genome? •Only 14 allozyme markers Social form completely associated to Gp-9 locus BB BB Bb Single queen form Multiple queen form (>15% )(< 5% ) x xx ✖ ✔ Locus! 0.3! 0.4! 0.5! 0.6! 0.7! 0.8! 0.9! 1.0! Single queen!Multiple queen! Est-6!Est-4! G 3pdh-1!C a-4!Pgm -4!Ddh-1!Pro-5! Pgm -3!Acoh-5!acoh-1!A cy-1!Pgm -1!Aat-2!Gp-9!
  40. 40. Sex chromosomes X Y Gp-9 B Gp-9 b SB Sb ? 1.Why non-recombining? “Social chromosomes” = supergene 2.Are SB and Sb differentiated? 3.What are the differences?
  41. 41. SBSB SBSb Single queen form Multiple queen form SBSB SB Sb Single queen colony Multiple queen colony SBSB SB Sb Single queen colony Multiple queen colony Summary: Fire ants have two colony types Summary: this is determined by a pair of social chromosomes
  42. 42. Research themes • Biomedical approaches • International population genomics surveys • Monitoring via sequencing • Major social transitions » social chromosomes » convergence » eusociality, queen number, parasitism... • 100-fold intra-specific variation in lifespan • Strengths of selection • Candidate genes/pathway Pollinator health Genome evolution Social evolution Modern bioinformatics tools & approaches (some at https://wurmlab.github.io )
  43. 43. SequenceServer
  44. 44. “Can you BLAST this for me?” BLAST But: •convoluted interface •challenging on custom data Antgenomes.org SequenceServer BLAST made easy is the most commonly used tool: >100,000 citations
  45. 45. http://www.sequenceserver.com/ If no config file:Asks interactive setup questions. If needed: Downloads BLAST binaries If needed: Formats FASTA into BLAST database. 1. Installing gem install sequenceserver ### Launched SequenceServer at: http://0.0.0.0:4567 2. Launch sequenceserver Demo Anurag Priyam - @yeban
  46. 46. http://www.sequenceserver.com/ Anurag Priyam @yeban
  47. 47. http://www.sequenceserver.com/ Anurag Priyam @yeban
  48. 48. Bionode
  49. 49. Timewasters • Client vs server-side code. • Workflows stalling (data download, cluster queues…) • Fragmented efforts - having to learn additional languages for specific tools + project- specific needs Bionode BrunoVieira @bmpvieira
  50. 50. Philosophy for flexibility Modules should: •(also) work in the web browser (when possible) •(also) work in the command-line •support streaming input/output gittergitter join chatjoin chat http://bionode.io BrunoVieira @bmpvieira
  51. 51. Difficulty writing scalable, reproducible and complex bioinformatic pipelines. Solution: Node.js everywhereStreams var ncbi = require('bionode-ncbi') var tool = require('tool-stream') var through = require('through2') var fork1 = through.obj() var fork2 = through.obj() ncbi .search('sra', 'Solenopsis invicta') .pipe(fork1) .pipe(dat.reads) fork1 .pipe(tool.extractProperty('expxml.Biosample.id')) .pipe(ncbi.search('biosample')) .pipe(dat.samples) fork1 .pipe(tool.extractProperty('uid')) .pipe(ncbi.link('sra', 'pubmed')) Node/Bionode for complex pipelines @bmpvieira
  52. 52. #"Get"descriptions"for"papers"related"to"SRA"search! bionode!ncbi!search!sra!Solenopsis!invicta!|!! !!!!!!!!tool3stream!extractProperty!uid!|!! !!!!!!!!bionode!ncbi!link!sra!pubmed!|!! !!!!!!!!tool3stream!extractProperty!destUID!|!
 !!!!!!!!bionode!ncbi!search!pubmed #"Get"URL"of"Solenopsis"invicta"genome" bionode3ncbi!urls!assembly!Solenopsis!invicta!|!json|!grep!genomic.fna! !http://ftp.ncbi.nlm.nih.gov/genomes/all/GCA_000188075.1_Si_gnG/GCA_000188075.1_Si_gnG_genomic.fna.gz http://bionode.io in the terminal #"Get"all"FASTQ"of"Arthropod"short"reads" bionode3ncbi!download!sra!arthropoda!|!bionode3sra!fastq3dump!3 #"Get"all"GFF"of"bacterial"genome"annotations" bionode3ncbi!download!gff!bacteria! @bmpvieira
  53. 53. BrunoVieira @bmpvieira Philosophy for flexibility Modules should: •(also) work in the web browser (when possible) •(also) work in the command-line •support streaming input/output Modules: •decentralised management. •small - just do one thing well. •few strict rules, but some strong recommendations (style, interfaces etc). gittergitter join chatjoin chat
  54. 54. BrunoVieira @bmpvieira Contributors gittergitter join chatjoin chat YOU? BioJS for visualisation Bionode for data handling
  55. 55. Baby steps towards improved efficiency, robustness & reproducibility
  56. 56. Biology has changed.
  57. 57. BIG
  58. 58. Geoffrey Chang: Crystallographer • Beckman FoundationYoung Investigator Award • Presidential Early Career Award Journal of Molecular Biology (2003) Chang. Structure of MsbA from Vibrio cholera: a multidrug resistance ABC transporter homolog in a closed conformation. PNAS (2004) Ma & Chang. Structure of the multidrug resistance efflux transporter EmrE from Escherichia coli. Science (2005) Reyes & Chang. Structure of the ABC transporter MsbA in complex with ADP vanadate and lipopolysaccharide. Science (2005) Pornillos et al. X-ray structure of the EmrE multidrug transporter in complex with a substrate. Science (2001) Chang & Roth. Structure of MsbA from E. coli: a homolog of the multidrug resistance ATP binding cassette (ABC) transporters. Science (2001) Chang & Roth.
  59. 59. earch Institute in next year, in a cer- Chang received a Award rs, the young ated a apers ctures ded in into a Swiss per in bt on a group cience gated, scover ispro- mns of density m had ucture. d used energy from adenosine triphosphate to trans- port molecules across cell membranes. These so-called ABC transporters perform many determination was at the root o cess: “He has an incredible d ethic. He really pushed the fie of getting things to no one else had be Chang’s data are go but the faulty so everything off. Ironically, anoth doc in Rees’s lab, K exposed the mistake tember issue of Na now at the Swiss F ofTechnology in Zu the structure of anA calledSav1866from aureus. The structur cally—and unexpe ent from that of pulling up Sav186 MsbA from S. typh computer screen, L realized in minutes structurewasinvert the “hand” of a mol Flipping fiasco. The structures of MsbA (purple) and Sav1866 (green) overlap little (left) until MsbA is inverted (right). California.The next year, in a cer- e White House, Chang received a l Early Career Award ts and Engineers, the ghest honor for young . His lab generated a high-profile papers e molecular structures proteins embedded in nes. e dream turned into a In September, Swiss published a paper in cast serious doubt on a cture Chang’s group ed in a 2001 Science en he investigated, horrified to discover madedata-analysispro- ipped two columns of ng the electron-density which his team had final protein structure. ly, his group had used m to analyze data for port molecules across cell membranes. These so-called ABC transporters perform many cess: “He has an ethic. He really p of get no on Chan but t every Iro doc in expos temb now a ofTec the str called aureu cally— ent f pullin MsbA comp realiz struct the “h a cha Flipping fiasco. The structures of MsbA (purple) and Sav1866 (green) overlap little (left) until MsbA is inverted (right). Sav1866 Dawson & Locher (2006) Nature Science(2001)Chang&Roth.Science (2001) Chang & Roth. Comparison with 3D structure of ortholog Science (2001) Chang & Roth.
  60. 60. http://wurmlab.github.io LETTERS I BOOKS I POLICY FORUM I EDUCATION FORUM I PERSPECTIVES 1878 1880 1882 LETTERS edited by Etta Kavanagh Retraction WE WISH TO RETRACT OUR RESEARCH ARTICLE “STRUCTURE OF MsbA from E. coli:A homolog of the multidrug resistanceATP bind- ing cassette (ABC) transporters” and both of our Reports “Structure of the ABC transporter MsbA in complex with ADP•vanadate and lipopolysaccharide”and“X-raystructureoftheEmrEmultidrugtrans- porter in complex with a substrate” (1–3). The recently reported structure of Sav1866 (4) indicated that our MsbA structures (1, 2, 5) were incorrect in both the hand of the struc- ture and the topology. Thus, our biological interpretations based on these inverted models for MsbA are invalid. Anin-housedatareductionprogramintroducedachangeinsignfor anomalous differences.This program, which was not part of a conven- tional data processing package, converted the anomalous pairs (I+ and I-) to (F- and F+), thereby introducing a sign change. As the diffrac- tion data collected for each set of MsbA crystals and for the EmrE crystals were processed with the same program, the structures reported in (1–3, 5, 6) had the wrong hand. The error in the topology of the original MsbA structure was a con- sequence of the low resolution of the data as well as breaks in the elec- tron density for the connecting loop regions. Unfortunately, the use of the multicopy refinement procedure still allowed us to obtain reason- able refinement values for the wrong structures. The Protein Data Bank (PDB) files 1JSQ, 1PF4, and 1Z2R for MsbA and 1S7B and 2F2M for EmrE have been moved to the archive of obsolete PDB entries. The MsbA and EmrE structures will be recalculated from the original data using the proper sign for the anom- alous differences, and the new Ca coordinates and structure factors will be deposited. We very sincerely regret the confusion that these papers have caused and, in particular, subsequent research efforts that were unpro- ductive as a result of our original findings. GEOFFREY CHANG, CHRISTOPHER B. ROTH, CHRISTOPHER L. REYES, OWEN PORNILLOS, YEN-JU CHEN, ANDY P. CHEN Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. References 1. G. Chang, C. B. Roth, Science 293, 1793 (2001). 2. C. L. Reyes, G. Chang, Science 308, 1028 (2005). 3. O. Pornillos, Y.-J. Chen, A. P. Chen, G. Chang, Science 310, 1950 (2005). 4. R. J. Dawson, K. P. Locher, Nature 443, 180 (2006). 5. G. Chang, J. Mol. Biol. 330, 419 (2003). 6. C. Ma, G. Chang, Proc. Natl. Acad. Sci. U.S.A. 101, 2852 (2004). MsbA from E. coli:A homolog of the multidrug resistanceATP bind- ing cassette (ABC) transporters” and both of our Reports “Structure of the ABC transporter MsbA in complex with ADP•vanadate and lipopolysaccharide”and“X-raystructureoftheEmrEmultidrugtrans- porter in complex with a substrate” (1–3). The recently reported structure of Sav1866 (4) indicated that our MsbA structures (1, 2, 5) were incorrect in both the hand of the struc- ture and the topology. Thus, our biological interpretations based on these inverted models for MsbA are invalid. Anin-housedatareductionprogramintroducedachangeinsignfor anomalous differences.This program, which was not part of a conven- tional data processing package, converted the anomalous pairs (I+ and I-) to (F- and F+), thereby introducing a sign change. As the diffrac- tion data collected for each set of MsbA crystals and for the EmrE crystals were processed with the same program, the structures reported in (1–3, 5, 6) had the wrong hand. The error in the topology of the original MsbA structure was a con- sequence of the low resolution of the data as well as breaks in the elec- 1860 Untilrecently,GeoffreyChang’scareerwason a trajectory most young scientists only dream about. In 1999, at the age of 28, the protein crystallographer landed a faculty position at the prestigious Scripps Research Institute in San Diego, California.The next year, in a cer- emony at the White House, Chang received a Presidential Early Career Award for Scientists and Engineers, the country’s highest honor for young researchers. His lab generated a stream of high-profile papers detailing the molecular structures of important proteins embedded in cell membranes. Then the dream turned into a nightmare. In September, Swiss researchers published a paper in Nature that cast serious doubt on a protein structure Chang’s group had described in a 2001 Science paper. When he investigated, Chang was horrified to discover thatahomemadedata-analysispro- 2001 Science paper, which described the struc- tureofaproteincalledMsbA,isolatedfromthe bacterium Escherichia coli. MsbA belongs to a huge and ancient family of molecules that use energy from adenosine triphosphate to trans- port molecules across cell membranes. These so-called ABC transporters perform many Sciences and EmrE, a differ Crystalliz five membra was an incred postdoc advis nia Institute o proteins are a because they ously diffic needed for x- determination cess: “He has ethic. He real of no Ch bu ev do ex tem no of the cal au ca en pu A Scientist’s Nightmare: Software Problem Leads to Five Retractions SCIENTIFIC PUBLISHING
  61. 61. ! Geoffrey Chang • Beckman FoundationYoung Investigator Award • Presidential Early Career Award Science (2001) Chang & Roth. Structure of MsbA from E. coli: a homolog of the multidrug resistance ATP binding cassette (ABC) transporters. Journal of Molecular Biology (2003) Chang. Structure of MsbA from Vibrio cholera: a multidrug resistance ABC transporter homolog in a closed conformation. PNAS (2004) Ma & Chang. Structure of the multidrug resistance efflux transporter EmrE from Escherichia coli. Science (2005) Reyes & Chang. Structure of the ABC transporter MsbA in complex with ADP vanadate and lipopolysaccharide. Science (2005) Pornillos et al. X-ray structure of the EmrE multidrug transporter in complex with a substrate. 1860 Untilrecently,GeoffreyChang’scareerwason a trajectory most young scientists only dream about. In 1999, at the age of 28, the protein crystallographer landed a faculty position at the prestigious Scripps Research Institute in San Diego, California.The next year, in a cer- emony at the White House, Chang received a Presidential Early Career Award for Scientists and Engineers, the country’s highest honor for young researchers. His lab generated a stream of high-profile papers detailing the molecular structures of important proteins embedded in cell membranes. Then the dream turned into a nightmare. In September, Swiss researchers published a paper in Nature that cast serious doubt on a protein structure Chang’s group had described in a 2001 Science paper. When he investigated, Chang was horrified to discover thatahomemadedata-analysispro- 2001 Science paper, which described the struc- tureofaproteincalledMsbA,isolatedfromthe bacterium Escherichia coli. MsbA belongs to a huge and ancient family of molecules that use energy from adenosine triphosphate to trans- port molecules across cell membranes. These so-called ABC transporters perform many Sciences and EmrE, a differ Crystalliz five membra was an incred postdoc advis nia Institute o proteins are a because they ously diffic needed for x- determination cess: “He has ethic. He real of no Ch bu ev do ex tem no of the cal au ca en pu A Scientist’s Nightmare: Software Problem Leads to Five Retractions SCIENTIFIC PUBLISHING
  62. 62. http://wurmlab.github.io This is costly For: •the individual •collaborators •the institution •1000s of researchers performing follow-up work •science •society
  63. 63. http://genome.gov/sequencingcosts
  64. 64. http://wurmlab.github.io • Understanding/visualising/analysing/massaging big data is hard. • Biology/life is complex. • Biologists lack computational training. • Field is young. • Analysis tools (generally) suck: • badly written • badly tested • hard to install • output quality… often questionable. • Data sizes keep growing! • Data formats keep changing :( Genome bioinformatics is hardBiology is harder than (many) other data sciences
  65. 65. http://wurmlab.github.io Some sources of inspiration
  66. 66. http://wurmlab.github.io Community Page Best Practices for Scientific Computing Greg Wilson1 *, D. A. Aruliah2 , C. Titus Brown3 , Neil P. Chue Hong4 , Matt Davis5 , Richard T. Guy6¤ , Steven H. D. Haddock7 , Kathryn D. Huff8 , Ian M. Mitchell9 , Mark D. Plumbley10 , Ben Waugh11 , Ethan P. White12 , Paul Wilson13 1 Mozilla Foundation, Toronto, Ontario, Canada, 2 University of Ontario Institute of Technology, Oshawa, Ontario, Canada, 3 Michigan State University, East Lansing, Michigan, United States of America, 4 Software Sustainability Institute, Edinburgh, United Kingdom, 5 Space Telescope Science Institute, Baltimore, Maryland, United States of America, 6 University of Toronto, Toronto, Ontario, Canada, 7 Monterey Bay Aquarium Research Institute, Moss Landing, California, United States of America, 8 University of California Berkeley, Berkeley, California, United States of America, 9 University of British Columbia, Vancouver, British Columbia, Canada, 10 Queen Mary University of London, London, United Kingdom, 11 University College London, London, United Kingdom, 12 Utah State University, Logan, Utah, United States of America, 13 University of Wisconsin, Madison, Wisconsin, United States of America Introduction Scientists spend an increasing amount of time building and using software. However, most scientists are never taught how to do this efficiently. As a result, many are unaware of tools and practices that would allow them to write more reliable and maintainable code with less effort. We describe a set of best practices for scientific software development that have solid foundations in research and experience, and that improve scientists’ productivity and the reliability of their software. Software is as important to modern scientific research as telescopes and test tubes. From groups that work exclusively on computational problems, to traditional laboratory and field scientists, more and more of the daily operation of science revolves around developing new algorithms, managing and analyzing the large amounts of data that are generated in single research projects, combining disparate datasets to assess synthetic problems, and other computational tasks. Scientists typically develop their own software for these purposes because doing so requires substantial domain-specific knowledge. As a result, recent studies have found that scientists typically spend 30% or more of their time developing software [1,2]. However, 90% or more of them are primarily self-taught [1,2], and therefore lack exposure to basic software development practices such as writing maintainable code, using version control and issue error from another group’s code was not discovered until after publication [6]. As with bench experiments, not everything must be done to the most exacting standards; however, scientists need to be aware of best practices both to improve their own approaches and for reviewing computational work by others. This paper describes a set of practices that are easy to adopt and have proven effective in many research settings. Our recommenda- tions are based on several decades of collective experience both building scientific software and teaching computing to scientists [17,18], reports from many other groups [19–25], guidelines for commercial and open source software development [26,27], and on empirical studies of scientific computing [28–31] and software development in general (summarized in [32]). None of these practices will guarantee efficient, error-free software development, but used in concert they will reduce the number of errors in scientific software, make it easier to reuse, and save the authors of the software time and effort that can used for focusing on the underlying scientific questions. Our practices are summarized in Box 1; labels in the main text such as ‘‘(1a)’’ refer to items in that summary. For reasons of space, we do not discuss the equally important (but independent) issues of reproducible research, publication and citation of code and data, and open science. We do believe, however, that all of these will be much easier to implement if scientists have the skills we describe. Education A Quick Guide to Organizing Computational Biology Projects William Stafford Noble1,2 * 1 Department of Genome Sciences, School of Medicine, University of Washington, Seattle, Washington, United States of America, 2 Department of Computer Science and Engineering, University of Washington, Seattle, Washington, United States of America Introduction Most bioinformatics coursework focus- es on algorithms, with perhaps some components devoted to learning pro- gramming skills and learning how to use existing bioinformatics software. Un- fortunately, for students who are prepar- ing for a research career, this type of curriculum fails to address many of the day-to-day organizational challenges as- sociated with performing computational experiments. In practice, the principles behind organizing and documenting computational experiments are often learned on the fly, and this learning is strongly influenced by personal predilec- tions as well as by chance interactions with collaborators or colleagues. The purpose of this article is to describe one good strategy for carrying out com- putational experiments. I will not describe profound issues such as how to formulate hypotheses, design experiments, or draw conclusions. Rather, I will focus on relatively mundane issues such as organiz- ing files and directories and documenting understanding your work or who may be evaluating your research skills. Most com- monly, however, that ‘‘someone’’ is you. A few months from now, you may not remember what you were up to when you created a particular set of files, or you may not remember what conclusions you drew. You will either have to then spend time reconstructing your previous experiments or lose whatever insights you gained from those experiments. This leads to the second principle, which is actually more like a version of Murphy’s Law: Everything you do, you will probably have to do over again. Inevitably, you will discover some flaw in your initial preparation of the data being analyzed, or you will get access to new data, or you will decide that your param- eterization of a particular model was not broad enough. This means that the experiment you did last week, or even the set of experiments you’ve been work- ing on over the past month, will probably need to be redone. If you have organized and documented your work clearly, then repeating the experiment with the new under a common root directory. The exception to this rule is source code or scripts that are used in multiple projects Each such program might have a projec directory of its own. Within a given project, I use a top-leve organization that is logical, with chrono logical organization at the next level, and logical organization below that. A sample project, called msms, is shown in Figure 1 At the root of most of my projects, I have a data directory for storing fixed data sets, a results directory for tracking computa tional experiments peformed on that data a doc directory with one subdirectory per manuscript, and directories such as src for source code and bin for compiled binaries or scripts. Within the data and results directo ries, it is often tempting to apply a similar logical organization. For example, you may have two or three data sets agains which you plan to benchmark your algorithms, so you could create one directory for each of them under data In my experience, this approach is risky because the logical structure of your fina
  67. 67. http://software.ac.uk
  68. 68. http://wurmlab.github.io Specific Approaches/Tools 1. Write code for humans
  69. 69. http://wurmlab.github.io Write code for humans (not computers!) • For • yourself • colleagues / collaborators • reviewers • other random people who may reuse/improve your code • Respect conventions (e.g., a style guide)
  70. 70. te Damian ConwayUse whitespace/indentation! e Damian Conway Same information
  71. 71. Line length Strive to limit your code to 80 characters per line. This fits comfortably on a printed page with a reasonably sized font. If you find yourself running out of room, this is a good indication that you should encapsulate some of the work in a separate function. ant_measurements <- read.table(file = '~/Downloads/Web/ant_measurements.txt', header=TRUE, se ant_measurements <- read.table(file = '~/Downloads/Web/ant_measurements.txt', header = TRUE, sep = 't', col.names = c('colony', 'individual', 'headwidth', 'mass') ) ant_measurements <- read.table(file = '~/Downloads/Web/ant_measurements.txt', header=TRUE, 
 sep='t', col.names = c('colony', 'individual', 'headwidth', 'mass')) R style guide extract http://r-pkgs.had.co.nz/style.html
  72. 72. R style guide extract http://r-pkgs.had.co.nz/style.html
  73. 73. http://wurmlab.github.io
  74. 74. http://wurmlab.github.io Write code for humans (not computers!) • For • yourself • colleagues / collaborators • reviewers • other random people who may want to reuse your code • Respect conventions (e.g., a style guide) • Don't optimise (generally…)
  75. 75. http://wurmlab.github.io Code reviews: ask a peer to (critically) read your analysis code. Or do peer-programming sessions
  76. 76. http://wurmlab.github.io Specific Approaches/Tools 1. Write code for humans 2. Organise mindfully
  77. 77. Eliminate redundancy DRY: Don’t RepeatYourself & don't reinvent the wheel. Organise mindfully
  78. 78. http://wurmlab.github.io Organise mindfully http://bit.ly/projectstruct Choose a standard/ template and stick to it! Choose a standard/ template and stick to it!
  79. 79. Organise mindfully http://bit.ly/projectstruct
  80. 80. http://wurmlab.github.io Specific Approaches/Tools 1. Write code for humans 2. Organise mindfully 3. Plan for mistakes
  81. 81. Automatically check consistency with style guide install.packages("lint") # once library(lint) # everytime lint("file_to_check.R")
  82. 82. http://wurmlab.github.io Create code tests that are easy to run • Unit tests == checking edge cases to see if the function works # do your stuff # e.g. define speed() function library(testthat) expect_that(speed(km = 0, minutes = 60), equals(0)) expect_that(speed(km = 60, minutes = 60), equals(1)) expect_that(speed(km = -4, minutes = 60), throws_error()) expect_that(nrow(significant_SNPs), 42) expect_that(my_model, is_a("lm")) • Integration tests == "full analysis" but on small data with known results • e.g. on fakeVCF genotype file of 2 loci (one true positive, one true negative) • Add sanity checks. E.g. the following should fail rather than return something incorrect.
 speed(km= "twenty", minutes=20) speed(km = -4, minutes = 60)
  83. 83. http://wurmlab.github.io "Continuous integration": Tests should run automagically. So you don't have to remember (or find time) to do it. "http://github.org Tests run automatically http://travis-ci.org If unexpected result: #
  84. 84. http://wurmlab.github.io Specific Approaches/Tools 1. Write code for humans 2. Organise mindfully 3. Plan for mistakes 4. Use tools that reduce risks
  85. 85. http://wurmlab.github.io Use tools that reduce risks • Ensure computers are set up for productivity. E.g.,: • use GNU parallel on an 80-core machine when more appropriate than submitting to queue • If you need to make a "pipeline", use software designed for this. E.g.: • Snakemake • Nextflow • (etc) • too many examples to discuss here
  86. 86. knitr/rmarkdown/ jupyter Analysis & report in one. analysis.Rmd A minimal R Markdown example I know the value of pi is 3.1416, and 2 times pi is 6.2832. To c library(knitr); knit( minimal.Rmd ) A paragraph here. A code chunk below: 1+1 ## [1] 2 .4-.7+.3 # what? it is not zero! ## [1] 5.551e-17 Graphics work too library(ggplot2) qplot(speed, dist, data = cars) + geom_smooth() ● ● ● ● ● ● ● ● ● ● ● ● ●●● ● ●● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ●●● ● ● 0 40 80 120 5 10 15 20 speed dist Figure 1: A scatterplot of cars
  87. 87. How to get users to adopt good practices? • Carrot (dual-benefit): • Use their motivation to have an easier life. "their motivation is the database" 
 "they see it, they understand it" -Thomasz? on SEEK • Piggyback off that so they do things better ("by stealth" -Carol) 
 • Stick: • When you're reviewing publications/grants • Politics: • Encourage funders / journals to require good practices.
  88. 88. Summary • Ants are cool • Biology is hard • We need to handle data better
  89. 89. y.wurm@qmul.ac.uk @yannick__ https://wurmlab.github.io @ Queen Mary U London Rodrigo Pracana Anurag Priyam @yeban Eckart Stolle Bruno Vieira @bmpvieira R Nichols & sbcsEvolve R Christie & T King / ITSR Apocrita Laurent Keller lab @ Lausanne J Wang, D Shoemaker,O Riba-Grognuz, M Nipitwattanaphon Ioannis Xenarios @ SIB DeWayne Shoemaker @ USDA Thanks!

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