mRNA-sequenciing for detection of heat-regulated alternative splicing - SRA

SRX253977: mRNA-sequenciing for detection of heat-regulated alternative splicing
3 ILLUMINA (Illumina HiSeq 2000) runs: 8.6M spots, 1.3G bases, 661Mb downloads

Design: MID-mRNA sequencing was carried out as suggested by Illumina, Inc. The method was adopted from mRNA-seq library prep protocol with addition of MID barcodes to the ds cDNA fragments and volume adjustment. Using the mRNA-seq kit, poly-A RNA were purified from 10-15 g of total RNA by oligo-dT beads and subjected to cation-catalyzed fragmentation for four minutes at 94˚C. The mRNA fragments were then converted into ds cDNA by random priming, and the ends were repaired and A-tailed. Multiplexing barcodes were then added to the DNA fragment ends with modifications from the paired-end gDNA library prep kit (Illumina, USA). The ligation products were size-selected on agarose gel (200-400 bp), subjected to 18 cycles of PCR and cleaned up by Ampure beads (Beckman Agencourt, USA). The libraries were quantified fluorometrically by Qubit (Invitrogen, USA) and BioAnalyzer High Sensitivity DNA chip (Agilent, USA). Mixing of the differentially barcoded samples for each lane was based on qPCR-derived relative concentration for amplifiable fraction of each library. This is the first repeat of sequencing, which uses a single-ended method to generate 75-bp reads. C, control; 1A, 1st HS; 3A, 2nd HS.

Submitted by: INSTITUTE OF PLANT AND MICROBIAL BIOLOGY, ACADEMIA SINICA, TAIPEI, TAIWAN (INSTITUTE OF PLANT AND MICROBIAL BIOLOGY, ACADEMIA)

Study: Physcomitrella patens Transcriptome or Gene expression

PRJNA192876 • SRP019809 • All experiments • All runs

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We used heat shock (HS) treatments and high-throughput mRNA sequencing to obtain HS transcriptomes in the moss Physcomitrella patens. Besides revealing differential gene expression at the transcriptional level, a transcriptome survey identified abundant AS events in the single-cell–type, undifferentiated, moss protonemal cell.

Sample: General Sample for Physcomitrella patens

SAMN01942529 • SRS402930 • All experiments • All runs

Organism: Physcomitrella patens

Library:

Name: Physco_heat

Instrument: Illumina HiSeq 2000

Strategy: RNA-Seq

Source: TRANSCRIPTOMIC

Selection: cDNA

Layout: PAIRED

Spot descriptor:

1 forward76 reverse

Runs: 3 runs, 8.6M spots, 1.3G bases, 661Mb

Run	# of Spots	# of Bases	Size	Published
SRR790623	2,575,951	386.4M	199.4Mb	2013-06-12
SRR790624	3,197,490	479.6M	258.7Mb	2013-06-12
SRR790625	2,833,797	425.1M	202.9Mb	2013-06-12

ID:: 354873

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