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| Status |
Public on May 11, 2017 |
| Title |
Sample 2 5-month |
| Sample type |
SRA |
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| Source name |
dermal fibroblast cells
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| Organism |
Bos taurus |
| Characteristics |
cell type: primary dermal fibroblasts
passage: 6
breed: Holstein
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| Treatment protocol |
No treatment
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| Growth protocol |
Primary dermal fibroblasts were revived from cryo-preservation simultaneously and cultured in DMEM, 1X ITS, 1X Pen/Strep antibiotic, 5% FBS into a T-75 until confluency
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted from cell lysate using 5 Prime PerfectPure DNA Kit per manufacturer's instructions.
Libraries were generated at Zymo Research with 2 ug of genomic DNA. Briefly, DNA was treated with MSPI, end-repaired, A-tailed and purified with Zymo's DNA clean and concentrator kit. Illumina adaptors were added per Illumina protocols. DNA was then bisulfite converted, PCR amplified and purified. Size selection was performed with a 4% NuSieve 3:1 agarose gel and libraries extracted with the Zymoclean Gel DNA Recovery Kit. DNA libraries were sequenced on the Illumina HiSeq2000 generating 50 bp paired end reads.
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
Reduced Representation |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
young
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| Data processing |
Illumina Casava software was used for basecalling.
A propriety pipeline by Zymo Research Inc. was used for bioinformatic analysis which assesses quality with FastQC (v0.11.1) and TrimGalore (v0.3.7) Illumina adaptors and bases with a Phred score less than 20 were trimmed along with the first 2 bases from the 3’ end of the sequence to avoid the final filled in cytosine being used to make a methylation call.
Alignment and methylation calling were performed with the Babraham Bismark software (v0.13.1). Bismark is designed for aligning bisulfite sequencing data while simultaneously making methylation calls. The result of bisulfite sequencing is four sets of potential sequences at any given locus. To determine which of these four potential sequences is correct, Bismark creates two bisulfite converted reference genomes, one that is a C to T conversion and one that is a G to A conversion to account for conversion on the reverse strand. Each sequence read is also bisulfite converted in silico and is aligned to the pre-converted version of the reference genome. The best alignment is then identified and used to make a methylation call. Alignments that are uniquely mapped were kept for analysis. Alignments that mapped to multiple regions were discarded.
Genome_build: BosTau8
Supplementary_files_format_and_content: bigbed
Supplementary_files_format_and_content: CpG_result_table.txt: text, all results coverage >5X
Supplementary_files_format_and_content: CpG_p_less_than_0.05.csv: Excel, all significant results, P<0.05, >5X coverage
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| Submission date |
Dec 09, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Filiz Korkmaz |
| E-mail(s) |
filiz.korkmaz@umassmed.edu
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| Phone |
413-885-6517
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| Organization name |
UMass Chan Medical School
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| Street address |
364 Plantation St.
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| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
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| Platform ID |
GPL15749 |
| Series (1) |
| GSE91088 |
DNA methylation analysis of fibroblasts isolated from 5- and 16-month Holstein cows |
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| Relations |
| BioSample |
SAMN06128095 |
| SRA |
SRX2409137 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2420838_zr1099_2A_CpG_meth.bigbed |
63.2 Mb |
(ftp)(http) |
BIGBED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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