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Sample GSM1111648

Query DataSets for GSM1111648

Status

Public on Jul 22, 2013

Title

160-J_RNA_c0_2

Sample type

SRA

Source name

Milk Fat Layer

Organism

Homo sapiens

Characteristics

subject id: 160
lactation stage: Transitional
extraction protocol: Hard spin, Washed once

Extracted molecule

polyA RNA

Extraction protocol

Soft Spin: Whole milk samples were centrifuged at 300 g for 10 min at 4 degrees C. The milk fat layers from three, 2.0 mL aliquots were transferred into a single new tube to which 500 µL TRIzol® was added and stored at -80 degrees C
Hard Spin: Whole milk samples were centrifuged at 15000 g for 10 min at 4 degrees C. The milk fat layers from three, 2.0 mL aliquots were transferred into a single new tube to which 500 µL TRIzol® was added and stored at -80 degrees C
Wash: After a hard spin at a warmer temperature, (15,000 g at 12 degrees C) the milk fat layers were transferred to a clean set of 3 tubes and re-suspended in 1 mL PBS+10µM EDTA. After centrifuging again (15,000 g at 12 degrees C), the milk fat layers from all 3 tubes were transferred to a single tube, re-suspended in 1 mL PBS, and centrifuged a final time (15,000 g at 12 degrees C). The remaining milk fat layer was transferred into a single new tube to which 500 µL TRIzol® was added and stored at -80 degrees C.
RNA Isolation: Total RNA was extracted and purified in batches of 16 using the PROMEGA Maxwell 16™ integrated system (Promega Corporation, Madison, WI).
RNA libraries were prepared for sequencing using the Illumina TruSeq RNA Kit and standard Illumina Protocols

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Data processing

Basecalls performed using Illumina software
Reads mapped to the genome using TopHat with these options: tophat -G hg19.gtf -o tophat -p 4 -r 0
Mapped reads were indexed using samtools: samtools index tophat/accepted_hits.bam
Normalized transcript abundances determined using Cufflinks: cufflinks -u -b hg19.fa -G hg19.gtf -p 4 -o cufflinks tophat/accepted_hits.bam
Genome_build: hg19
Supplementary_files_format_and_content: Compressed tab-delimited text files contain transcript abundance measurements (FPKM) summarized at the gene level

Submission date

Apr 01, 2013

Last update date

May 15, 2019

Contact name

Danielle G Lemay

E-mail(s)

dglemay@ucdavis.edu

Organization name

University of California Davis

Department

Genome Center