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Overview Of Array Based Copy Number
 

Overview Of Array Based Copy Number

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    Overview Of Array Based Copy Number Overview Of Array Based Copy Number Document Transcript

    •                       Overview of array-based Copy Number analysis for Cytogenetics   Topics in Copy Number variation analysis       October  2009   ©2009  infoQuant,  Ltd   1   Doc:  OCPEQS1  
    •     Contents   Overview  .................................................................................................................................................................................  2   Tools  ........................................................................................................................................................................................  2   An  array-­‐based  Cytogenetic  Test  ............................................................................................................................................  2   Using  parental  samples  ...........................................................................................................................................................  4   Utilizing  accumulated  patient  data  .........................................................................................................................................  4   Gene  significance  ....................................................................................................................................................................  6   Conclusion  ...............................................................................................................................................................................  8     Overview Array  CGH  technique  is  going  through  a  transition  from  being  purely  a  research  tool  to  being  used  in  clinical  Cytogenetic   tests  on  a  daily  basis.  Such  transition  calls  for  a  new  generation  of  analytical  solutions  capable  of  semi-­‐automated  and   reliable  copy  number  reporting.   Tools oneClickCGH®  is  a  unique  software  package  that  facilitates  automated  analysis  of  array  CGH  data  and  user-­‐friendly  copy   number  reporting  for  individual  samples.  Cytogenetic  labs  can  use  this  package  to  support  identification  of  clinically   relevant  copy  number  changes  and  to  build  robust  and  comprehensive  reports.  It  is  capable  of  analyzing  medium-­‐  and   high-­‐resolution  data  from  various  array  CGH  and  SNP  platforms.  Statistical  algorithms  used  in  this  software  ensure   reliability  of  aberration  detection,  which  assists  Cytogenetic  users  in  producing  their  final  report  documents.   CGH  Fusion®  is  a  leading  commercially  validated  package  used  for  analysis  of  array  CGH  data  across  multiple  samples.  It   is  capable  of  building  copy  number  profiles  for  large  sets  of  samples  within  a  specific  biological  context.  CGH  Fusion  can   facilitate  constriction  of  libraries  of  copy  number  profiles  characterizing  specific  disorders  or  cohorts  of  healthy  patients   in  a  simple  user-­‐friendly  manner.  It  can  also  provide  valuable  biological  insights  into  regions  of  frequent  copy  number   change  by  incorporating  various  gene  annotations  and  known  CNVs.   An array-based Cytogenetic Test Usually  analysis  of  test  data  extracted  with  array  CGH  or  SNP  array  starts  with  detection  of  suspected  regions  of  copy   number  changes.  Several  basic  operations  are  performed  on  raw  data  (depending  on  the  actual  array  platform  used):   background  subtraction,  normalization,  filtering  of  outlier  replicates  and  actual  detection  of  regions  of  abnormal  log-­‐ ©2009  infoQuant,  Ltd   2   Doc:  OCPEQS1  
    •     ratio  values.  All  aforementioned  operations  can  be  performed  automatically  and  almost  no  manual  intervention  is   needed  at  that  stage.      However,  not  all  automatically  detected  chromosomal  aberration  regions  need  to  be  reported  as  relevant  to  the  test.   Some  of  them  can  be  simply  experimental  defects  and  some  come  from  regions  of  known  genetic  variance,  which  do  not   contribute  to  the  clinical  condition  under  study.     Therefore  a  manual  procedure  of  aberration  approval  and  reporting  is  needed.  Such  procedure  usually  is  centered   around  building  a  final  report  with  manually  approved  copy  number  changes  based  on  references  to  various  knowledge   databases  (public  or  internal).  Such  aberration  reporting  procedure  can  be  streamlined  with  oneClickCGH,  which  allows   creation  of  comprehensive  report  documents  in  various  formats  and  facilitates  references  to  publicly  available   databases  such  as  the  Data  Base  of  Genomic  Variants,  UCSC  Genome  Browser,  OMIM,  Gene  Ontology  database  etc.   First,  all  detected  copy  number  changes  need  to  be  reviewed  and  gene  and/or  known  CNV  annotations  can  be  used  to        After  all  detected  aberrations  have  been  approved  and  necessary  comments  have  been  added,  report  section  of   oneClickCGH  contains  only  relevant  regions  of  copy  number  change  that  can  be  saved  as  a  report  document     ©2009  infoQuant,  Ltd   3   Doc:  OCPEQS1  
    •     Using parental samples ones.     ical.  However,     when  such  data  are  available,  CGH  Fusion can  provide  a  user-­‐friendly  environment  for  identification  and  reporting     User  can  simply  load  all  available  samples  into  CGH  Fusion,  analyze  them  simultaneously  and  review  them  side  by  side.   Plot  below  illustrates  this  operation.     It  is  easy  to  see  from  this  plot  that  two  out  of  three   mother s)  demonstrate  similar  aberrations,   which  also  coincide  with  a  region  of  known  CNVs  from  Toronto  DGV  highlighted  with  orange  at  the  top  of  the  plot.   a  known  CNV  inherited  from  one  of   parent  genomes.     Utilizing accumulated patient data Ultimately,  every  diagnostician  would  like  to  be  able  to  understand  clinical  relevance  of  a  chromosomal  anomaly  under   scope  by  comparing  frequency  of  its  occurrence  in  patients  with  a  particular  disorder  vs.  healthy  patients.  This  can  be   ©2009  infoQuant,  Ltd   4   Doc:  OCPEQS1  
    •     done  if  array  CGH  data  for  large  cohorts  of  patients  have  been  accumulated  by  the  lab.  If  sufficient  number  of  patients   with  the  disorder  were  tested,  such  data  can  be  used  to  build  an  aberration  frequency  profile  for  the  disease.  On  the   other  hand,  you  can  use  data  from  healthy  patients  to  assess  frequency  of  normal  CNV  occurrence,  which  may  explain   some  of  the  anomalies  initially  thought  to  be  related  to  the  disorder  under  study.   CGH  Fusion  can  help  users  build  a  comprehensive  and  informative  aberration  frequency  profile  for  every  patient  group   that  lab  has  accumulated  array  Copy  Number  data  for.  Publicly  available  datasets  can  also  be  used  for  such  purpose.     You  can  easily  load  hundreds  or  thousands  of  samples  into  CGH  Fusion  and  perform  batch  copy  number  identification  on   them.  Output  of  that  procedure  can  then  be  used  to  build  an  aberration  frequency  profile  illustrated  below.     Such  profile  clearly  indicates  regions  of  frequent  copy  number  anomalies  for  the  patient  cohort  under  study  and   provides  a  quantitative  measure  of  aberration  frequency  in  the  cohort.   By  re-­‐using  profiles  constructed  with  CGH  Fusion  in  oneClickCGH  during  a  routine  Cytogenetic  test,  one  can   quantitatively  estimate  probability  of  an  aberration  region  contributing  to  a  particular  disorder.   ©2009  infoQuant,  Ltd   5   Doc:  OCPEQS1  
    •       Illustration  above  demonstrates  how  a  gain  and  a  loss  detected  in  a  sample  (upper  plot)  coincide  with  regions  of  high   CNV  frequency  highlighted  by  green  and  red  peaks  of  aberration  frequency  profile  for  healthy  individuals  (lower  plot).   Aberration  frequency  profile  was  constructed  using  data  from  HapMap  samples  extracted  using  Affymetrix  SNP  6.0  chip.   Gene significance Finally,  gene  annotations  for  a  chromosomal  region  of  interest  and  any  biological  information  associated  with  those  are   probably  the  most  informative  pieces  of  the  puzzle  when  it  comes  to  interpreting  detected  aberrations.  Of  course,  one   can  always  manually  review  sets  of  genes  from  suspect  regions  of  copy  number  change  and  skim  through  various   publications  and  biological  information  associated  with  those  genes.  This  procedure  can  be  semi-­‐automated  with   oneClickCGH  or  CGH  Fusion,  where  gene  annotations  are  conveniently  located  alongside  log-­‐ratio  plot.     Picture  below  demonstrates  how  users  can  review  genes  within  a  region  of  interest  and  identify  conditions  genes  under   scope  have  been  associated  with  in  available  publications.  This  example  illustrates  how  Angelman  syndrome  can  be   added  to  the  list  of  conditions  a  copy  number  loss  on  chromosome  15  may  be  related  to.     ©2009  infoQuant,  Ltd   6   Doc:  OCPEQS1  
    •       This  procedure  can  be  further  automated  by  short-­‐listing  genes  affected  by  copy  number  changes  along  genome  for   current  sample.  By  a  click  of  a  button  users  can  display  a  table  listing  affected  genes  and  corresponding  significance  p-­‐ values  ordered  from  most  statistically  significant  to  least  statistically  significant.  The  table  also  lists  conditions  cited  in   association  with  the  genes  and  corresponding  Ontology  terms.  Below  you  can  see  how  such  list  also  highlights  gene   UBE3A  identified  in  the  previous  paragraph.   ©2009  infoQuant,  Ltd   7   Doc:  OCPEQS1  
    •       Conclusion oneClickCGH  in  combination  with  CGH  Fusion  provide  a  powerful  suite  of  DNA  Copy  Number  analysis  tools  for  array   users.  These  software  products  provide  the  new  level  of  in-­‐depth  array  data  interpretation  and  comprehensive  Copy   Number  reporting  that  can  be  beneficial  for  an  individual  user  and  for  the  laboratory  workflow  as  a  whole.  It  is  a  truly   universal  software  suite  that  is  capable  of  handling  array  data  of  any  resolution  obtained  with  any  hardware  platform:   Affymetrix,  Agilent,  Illumina,  PerkinElmer,  Roche-­‐Nimblegen  and  more.         ©2009  infoQuant,  Ltd   8   Doc:  OCPEQS1  
    •       ©2009  infoQuant,  Ltd   9   Doc:  OCPEQS1