Athurva Gore/LabNotes/2010-1-11: Difference between revisions

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=Padlock Probe Software Modifications=
=Padlock Probe Software Modifications=
* Implement ViennaRNA algorithm for folding energy instead (Free for commercial use, same basic principles)
* Implement ViennaRNA algorithm for folding energy instead (Free for commercial use, same basic principles)
* Finish patent and IP form (already did copyright a while back)
** Copied a lot from copyright form.
** Can finish it out tonight, get to Dr. Zhang by tomorrow


=IPS Paper=
=IPS Paper=

Latest revision as of 03:09, 15 January 2010

Padlock Probe Software Modifications[edit]

  • Implement ViennaRNA algorithm for folding energy instead (Free for commercial use, same basic principles)

IPS Paper[edit]

  • Waiting to hear back from Science.
  • If we hear back negatively, send Letter to Nature format paper to Dr. Zhang and Dr. Goldstein
    • Edit LtN form so it sounds even better...sounds ok to me, but want it to be best quality

Digital Quantification[edit]

  • Data Analysis
  • LOOK AT CONNECTIVITY DATABASE

PGP Samples[edit]

  • Calculated our exomic coverage at depth 8 and quality 30 for PGP1, PGP2, and PGP4
    • SNPs found were all 94-95% in dbSNP. This means that we can probably trust the data.
  • Need to fill in below table, see what our approximate optimal amount of sequencing is.
  • Currently:
    • Base-calling PGP7
    • Mapping PGP6
    • Mapping PGP10
    • PGP8 and PGP9 still remain
  • Need to complete mapping for PGP 6-10.
Sample Reads Estimated Usable Reads CCDS Coverage (depth 8, quality 30)
PGP1 73,656,659 58,630,458 22,543,941 (78%)
PGP2 13,777,534 8,325,274 9,964,899 (35%)
PGP3 19,642,850 12,613,974 11,716,319 (40%)
PGP4 39,022,194 21,228,150 17,859,704 (62%)
PGP5 18,620,501 12,083,551 11,260,366 (39%)
PGP6 44,542,386 23,913,315
PGP7 28,619,340 17,384,463
PGP8 116,237,040 63,690,701
PGP9 117,277,366 60,695,633
PGP10 26,108,372 13,478,483

Lab Meeting[edit]

  • Dr. Guanghui Liu, from Belmonte Lab in Salk
    • Reprogramming Lab

Potential Role of Nuclear Reprogramming

Reprogramming and iPS iPS cells - makes derivation of patient-specific stem cells possible Slow (2-3 weeks), frequency low (0.1%)

Several factors can interfere with or promote reprogramming p53, p21, p16, p19, telomere shortening (senesence) - inhibitory ERK, TGF-b, Wnt (signaling pathways) - can be either inhibitory or excitatory cell division genes - excitatory

Nuclear envelope - role in reprogramming

Lamin/AC disappears in stem cells - causes a plastic/pliable nuclear envelope Somatic nuclear envelope is stiff due to presence of Lamin A/C

Nuclear periphery - related to gene silencing

Can nuclear envelope/lamina regulate reprogramming?

Laminopathies - mutation in Lamin A has been linked to diseases

HGPS (Progeria) is caused by a SNP that prevents alternative splicing in Lamin A Premature aging Nuclear envelope is deformed

HGPS cells should be hard to reprogram, theoretically Highly aged cells DNA damage in cells, p53 is activated Shortened telomere Cells grow slowly

Reprogramming of BJ and HGPS cells HGPS cells were resistant to viral infection due to nuclear envelope changes However, colony structure forms in HGPS cells more readily than in BJ cells! Nanog staining shows that HGPS cells have 10-20 fold better reprogramming efficiency!

Lamin A and C is gone in iPS cells Lamin B expression is enhanced

Test drugs on normal fibroblasts Can we use anti-lamin drugs to enhance reprogramming efficiency? Tested with FTI-277 Colonies more readily form! Improved reprogramming efficiency

Hypothesis HGPS nuclei are more readily epigenetically changed

HGPS have abnormal epigenetic markers

shRNA to knockdown of HP1 and SUV39H1 Mimic effect