User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/26: Difference between revisions
From OpenWetWare
(4 intermediate revisions by the same user not shown) | |||
Line 11: | Line 11: | ||
<!-- Precede finished items with a checkmark ✓ --> | <!-- Precede finished items with a checkmark ✓ --> | ||
* ChIP-seq: use input results to determine enrichment of U2OS H3K27me3 signals with Seqman NGen | * ChIP-seq: use input results to determine enrichment of U2OS H3K27me3 signals with Seqman NGen | ||
* TRIzol prep: U2OS PcTF+ & mock cells | |||
Line 37: | Line 38: | ||
* Step 3, 4 - Add Experiments to Import: added '''6_aln_sorted.bam''' (H3K27me3 ChIP), and '''inputU2OS.bam''' (DNA purified from unenriched chromatin) | * Step 3, 4 - Add Experiments to Import: added '''6_aln_sorted.bam''' (H3K27me3 ChIP), and '''inputU2OS.bam''' (DNA purified from unenriched chromatin) | ||
* Step 5 - Create binding proteins: created "H3K27me3" for 6_aln_sorted. | * Step 5 - Create binding proteins: created "H3K27me3" for 6_aln_sorted. | ||
* Set control inputU2OS as "yes" for "Is control?" | * Step 6 - Assigned H3K27me3 as protein label for 6_aln_sorted; Set control inputU2OS as "yes" for "Is control?"; Set inputU2OS as control for 6_aln_sorted. | ||
* Step 11 - Chose BED files only (wigs take up lots of disk space) | |||
* Step 12 - Deselected "Directionality" | |||
* | |||
* | |||
---- | |||
-- | '''U2OS TRIzol prep'''<br> | ||
# +PcTF, plate 1-A | |||
# +PcTF, plate 1-B | |||
# mock, plate 4-A | |||
# mock, plate 4-B | |||
(Should provide more than enough RNA. Discarded other cultures) | |||
* Cells were 100% confluent | |||
* Discarded growth medium | |||
* Added 2 mL TRIzol directly to plates | |||
* Incubated at r.t. for ~5 min. | |||
* Collected lysed cells from plate with gentle scraping (pipette tip) | |||
* Transferred 500 μL aliquots to 2.0 mL tubes. Stored at -80°C | |||
** 8x PcTF+ samples | |||
** 8x mock samples | |||
Revision as of 21:14, 27 June 2013
Pc-TF Genomics | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> |
06/26/13
ChIP seq analysis with Input control Array Star analysis - general instructions
H3K27me3 enrichment in U2OS
U2OS TRIzol prep
(Should provide more than enough RNA. Discarded other cultures)
|