Athurva Gore/LabNotes/2010-1-11
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