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| Status |
Public on Dec 12, 2022 |
| Title |
siCtrl_G9a ChIP-seq_Rep2 |
| Sample type |
SRA |
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| Source name |
Human prostate cancer cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: LNCaP
treatment/infection: siCtrl
antibody: G9a
vendor: bethyl laboratory
catalog number: A300-933A
lot/batch number: Lot 2
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| Treatment protocol |
LNCaP cells were infected with control shRNA (pLKO), shRNA targeting G9a (shG9a), shRNA targeting PALI1 (shPALI1) or/and shRNA targeting JARID2 (shJARID2), lentiviruses for 4-5 days. For siRNA-mediated interference, LNCaP cells were transfected with control siRNA (siCtrl) or G9a siRNA (siG9a) for 4 days.
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| Growth protocol |
LNCaP cells were maintained in RPMI supplemented with 10% FBS and 1% penicillin and streptomycin
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For SUZ12 and G9a ChIP-Seq, Chromatin in nuclear fraction were sheared to 200-500bp using an Covaris M220 Focused-ultrasonicator, Lysates were clarified from sonicated nuclei and protein-DNA complexes were immunoprecipitated using the indicated antibody. DNA was then reverse crosslinked from protein and purified. For H3K27me3, H3K9me2 CUT&Tag and PALI1 CUT&RUN, 0.5x10^6 LNCaP cells bound to Concanavalin A-coated beads were incubated with indicated primary antibodies and secondary antibody. pA-Tn5 was used to induce DNA tagmentation in CUT&Tag while pA-MNase was used to digest chromatin/DNA bound by the antibodies-protein complex in CUT&RUN, followed by Proteinase K digestion and PCI extraction.
For CUT&Tag, the ethanol precipitated DNA was immediately subjected to PCR amplification by Universal and barcoded i7 primer using NEBNext HiFi 2x PCR Master mix. The amplified library was purified by using Agencourt AMPure XP beads without size selection. For ChIP-Seq and CUT&RUN, ChIP-seq libraries were prepared from 3-5ng ChIPped DNA while CUT&RUN libraries were prepared from 15-20ng DNA using NEBNext® Ultra™ II DNA Library Prep Kit (NEB, E7645S), according to the manufacturer’s instructions. Postamplification libraries were size selected at 250–450 bp (ChIP-Seq) and 160-320bp (CUT&RUN) in length using Agencourt AMPure XP beads from Beckman Coulter and were quantified using the Library Quantification Kit from Illiumina (Kapa Biosystems, KK4603). All CUT&Tag, CUT&RUN and ChIP-Seq Libraries were pooled to a final concentration of 10nM and sequenced single-end using the Illumina HiSeq 4000.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiScanSQ |
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| Data processing |
Reads were aligned to the hg19 reference human genome using bowtie2
Peak-calling by HOMER v4.2
Genome_build: hg19
Assembly: hg19
Supplementary files format and content: bigWig files and peak excel sheet
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| Submission date |
Sep 02, 2022 |
| Last update date |
Dec 12, 2022 |
| Contact name |
Jindan Yu |
| E-mail(s) |
jindan.yu@emory.edu
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| Organization name |
Emory University
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| Department |
Urology
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| Lab |
Jindan Yu's lab
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| Street address |
E330, 1760 Haygood Dr NE
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| City |
Atlanta |
| State/province |
GA |
| ZIP/Postal code |
30322 |
| Country |
USA |
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| Platform ID |
GPL15456 |
| Series (2) |
| GSE175697 |
PALI1 Promotes Tumor Growth through Competitive Recruitment of PRC2 to G9A-target Chromatin for Dual Epigenetic Silencing |
| GSE212625 |
PALI1 Promotes Tumor Growth through Competitive Recruitment of PRC2 to G9A-target Chromatin for Dual Epigenetic Silencing [ChIP-seq] |
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| Relations |
| BioSample |
SAMN30651244 |
| SRA |
SRX17413110 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6541884_m636.bw |
87.5 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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