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DRX000815

FASTQ

SRA

Experiment Detail

TitleGSgt10019
Design Descriptionnone provided
OrganismMus musculus

Library Description

NameGSgt10019
StrategyAMPLICON
SourceTRANSCRIPTOMIC
SelectionRT-PCR
LayoutSINGLE
Construction ProtocolRNA was extracted using Isogen protocol and cDNA synthesized using the SuperScript II kit (Invitrogen) with oligo-dT primers (Invitrogen) according to manufacturer?s protocol. cDNAs were then amplified using ExTaq polymerase kit (TAKARA). TRAV14 (CCAATCTCTGACAGTCTGGGAA) and TRAC (AACTGGTACACAGCAGGTTCTG) primers were used for TRAV14 repertoire; TRBV2 (CGAACAGTATCTAGGCCACAATGC) and TRBC (GCACACGAGGGTAGCCTTTTGTTT) primers for TRBV2 repertoire. PCR program was as follow: 4 min at 94 ?C; x cycles of 30 s at 94 ?C, 30 s at 62 ?C, 30 s at 72 ?C; 5 min at 72 ?C 5; hold at 4 ?C. The number of cycles was adapted to the initial number of cells (24 or 25 cycles for conventional T cells, 27 to 29 for regulatory T cells) as we observed the formation of heteroduplexes in saturated reactions. PCR products were sequenced on a Roche Genome Sequencer FLX Titanium system after adapter ligation following standard procedures.

Platform

PlatformLS454
Instrument Model454 GS FLX Titanium

Processing

PipeSection
Step Index
Prev Step Index
Program2.3 (090928_1418)
Version

Spot Information

Number of Reads per Spots0
Spot Length400

Read Spec

Read Index 0
Read Label
Read ClassTechnical Read
Read TypeAdapter
Base Coord1
Read Index 1
Read Label
Read ClassTechnical Read
Read TypePrimer
Base Coord0
Read Index 2
Read Label
Read ClassApplication Read
Read TypeOther
Base Coord0

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Submission DRA000428 FTP
Study DRP000436
Sample DRS000778
Run DRR001218 FASTQ SRA
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