Athurva Gore/LabNotes/2009-9-9: Difference between revisions
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=Exome Resequencing= | =Exome Resequencing= | ||
==PGP4, 6, and 10== | |||
* Dr. Zhang pointed out that PGP4, 6, and 10 used size-selected libraries | |||
* Wanted to see if using a 5 bp throwout range works well with these libraries | |||
* For all three, success! | |||
** 10% false call rate with PGP10 | |||
** 5% non-dbSNP rate with PGP4 and PGP6 | |||
* Can therefore use a 5bp trimming due to size-selection; as expected. | |||
* Compute statistics, insert here | |||
* '''RESULTS:''' | |||
** ''ADD HERE'' | |||
=IPS and Cancer= | =IPS and Cancer= | ||
==LOH Mutations== | |||
* Need to check LOH mutations; is the new variant the damaging one? Or the old one? | * Need to check LOH mutations; is the new variant the damaging one? Or the old one? | ||
** If nondamaging version is lost, this is not actually harmful to the cell... | ** If nondamaging version is lost, this is not actually harmful to the cell... | ||
** Modified SIFT input to have fibroblast first, IPS second | ** Modified SIFT input to have fibroblast first, IPS second | ||
** Can now easily check if damage is gained or lost. | ** Can now easily check if damage is gained or lost. | ||
* ALL DAMAGING MUTATIONS: The non-damaging variant was lost via LOH! | |||
** True for most (all but 3) tolerated mutations as well | |||
** Strange. Will need to investigate with Primers... | |||
===Foreskin Fibroblast data=== | |||
* Recall that foreskin fibroblast clustered alone...and clustered together only when penalty for heterozygous mismatch was removed. | |||
* Could fibroblast have a bunch of extra heterozygous calls for some reason? | |||
** Check some of the LOH SNPs. What is happening at these positions? | |||
*** Quality issues? | |||
*** One strand capturing well? | |||
*** Maybe still issues near the 3' end? Had to cut it down to 15 bp, recall... | |||
==PCR Primers for target Genes== | |||
* Need to use Primer3 to obtain PCR primers for regions of interest. | |||
** Have five regions of interest: 2 from GOH, 3 from LOH | |||
** Get book on DNA methods? Or talk to Alice or Ida about it | |||
** Pull out sequences of an optimal length around SNP plus flanking regions |
Latest revision as of 23:53, 9 September 2009
[edit]
Exome Resequencing[edit]
PGP4, 6, and 10[edit]
- Dr. Zhang pointed out that PGP4, 6, and 10 used size-selected libraries
- Wanted to see if using a 5 bp throwout range works well with these libraries
- For all three, success!
- 10% false call rate with PGP10
- 5% non-dbSNP rate with PGP4 and PGP6
- Can therefore use a 5bp trimming due to size-selection; as expected.
- Compute statistics, insert here
- RESULTS:
- ADD HERE
IPS and Cancer[edit]
LOH Mutations[edit]
- Need to check LOH mutations; is the new variant the damaging one? Or the old one?
- If nondamaging version is lost, this is not actually harmful to the cell...
- Modified SIFT input to have fibroblast first, IPS second
- Can now easily check if damage is gained or lost.
- ALL DAMAGING MUTATIONS: The non-damaging variant was lost via LOH!
- True for most (all but 3) tolerated mutations as well
- Strange. Will need to investigate with Primers...
Foreskin Fibroblast data[edit]
- Recall that foreskin fibroblast clustered alone...and clustered together only when penalty for heterozygous mismatch was removed.
- Could fibroblast have a bunch of extra heterozygous calls for some reason?
- Check some of the LOH SNPs. What is happening at these positions?
- Quality issues?
- One strand capturing well?
- Maybe still issues near the 3' end? Had to cut it down to 15 bp, recall...
- Check some of the LOH SNPs. What is happening at these positions?
PCR Primers for target Genes[edit]
- Need to use Primer3 to obtain PCR primers for regions of interest.
- Have five regions of interest: 2 from GOH, 3 from LOH
- Get book on DNA methods? Or talk to Alice or Ida about it
- Pull out sequences of an optimal length around SNP plus flanking regions