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Genetics presentation(suggested edits)


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Genetics presentation(suggested edits)

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  2. 2. Differential regulation of the foraging gene associated with task behaviors inharvester ants<br />By Jonathan Kahn, Andrew Hoadley, and Claire Spitzer <br />
  3. 3. Summary<br />This paper is a first time series analysis of foraging gene expression in harvester ants. It shows that the task-specific expression patterns of foraging are aligned with the circadian rhythm of the ants. This data underlines the importance that time-related expression studies are extremely important and thus can be useful in explaining mechanisms that influence behaviour; such as foraging. <br />
  4. 4. Background<br />The individual behavior of a harvester ant is one thatchanges over their lifetime; <br />Transition from nest workers to foragers as they age.<br />This social organization is highly implicated(?associated with?) by a genetic pathway that involves the cGMP-activated protein kinase gene, foraging. <br />
  5. 5. cGMP-activated Protein Kinase Gene<br /><ul><li>This gene is associated with behaviour in the harvester ant (also many other species-used as an example) and is conserved amoung species although the expression patterns and functions of the gene vary across taxa. </li></li></ul><li>Background Continued<br />Many species such as the honeybee and the fruit fly provide examples as to how important the foraging gene is for behaviour and implications on the species structure; such as division of labour. <br />The foraging gene becomes much more interesting to look at after the discovery of task-specific expression in alignment with the circadian clock(?).<br />
  6. 6. Background Continued <br />This experiment looked to compare the amino acid sequence of foraging across social insects and observed whether the differential regulation of this gene is directly associated with task-related behaviours. <br />
  7. 7. Methods<br />The major techniques utilized during this experiment include:<br />Deep freeze storage of samples<br />qPCR/RT-PCR<br />DNA sequencing/analysis<br />
  8. 8. Deep Freeze Storage<br />Upon collection of individual ants, specimens were immersed in liquid nitrogen and stored at -70°C until dissection<br />This step is essential for the accuracy of the study and preservation of target RNA<br />Deep freeze storage of samples ensures:<br />Desired expression of RNA given time of sampling<br />Prevention of degradation of “foraging” RNA<br />
  9. 9. qPCR<br />Otherwise known as quantitative PCR (also abbreviated RT-PCR)<br />This technique was utilized in order to quantify the expression of the 130bp foraging gene at given intervals for which mRNA was extracted<br />What is normal PCR?<br />
  10. 10. qPCR (Traditional PCR)<br />Once mRNA is extracted from the specimen, designed primers and DNApolare added to the extraction to create cDNA (complimentary DNA) of the target gene<br />With PCR (polymerase chain reaction), primers and a high-temperature optimum DNApol are added to the extraction above to create DNA<br />This mixture is then heated and cooled repeatedly to amplify the DNA<br />
  11. 11. qPCR (Traditional PCR)<br />
  12. 12. qPCR (Traditional PCR)<br />However, this technique usually takes 20-40 cycles and can be used as a semi-quantitative tool at best since attachment of primers to DNA is not consistent<br />So What is the difference between PCR and qPCR?<br />
  13. 13. qPCR<br />With qPCR, a fluorescent probe is used<br />As DNApol elongates the target gene during cycles, the probe fluoresces and this is detected by a machine<br />After multiple cycles, the machine’s log of relative fluorescence detection can be analyzed to obtain an accurate idea of the mRNA that was originally present<br />
  14. 14. qPCR<br />
  15. 15. qPCR<br />Traditional PCR reveals results at the end which are unhelpful in wanting to quantify gene expression<br />qPCR, however, provides real time results which may be traced backwards to yield the level of mRNA that was originally present, essential to the evaluation of forgaing expression at a given time for this experiment<br />
  16. 16. DNA Sequencing/Analysis<br />A machine reads the target DNA that was developed by using Sanger sequencing principles<br />Using ddNTPs (A, T, G, and C nucleosides) which both fluoresce (by differing color) and terminate elongation, the machine can assemble the overlapping fragments of newly synthesized DNA according to fluorescence to reveal the sequence<br />This is then further analyzed by inputting the sequence into a global database, found at<br />
  17. 17. Results<br />
  18. 18. Catalytic Domain<br />Effector<br />Terminal Domain<br />Foraging Protein Sequence<br />
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  20. 20. Workers<br />Foragers<br />