SAMD00189824
External Id
Laboratory of Gene Expression, Medical Research Institute, Tokyo Medical and Dental University
INSDC center name
2021-01-29T00:00:00Z
INSDC first public
2021-01-29T08:17:11Z
INSDC last update
DRS171751
INSDC secondary accession
live
INSDC status
Generic
NCBI submission model
Generic.1.0
NCBI submission package
DRS171751
SRA accession
Synchronized L1 larvae of KH1668: smg-2 (yb979) I were fed with an E. coli strain OP50 in S-complete medium for 3 hours at 20 degree Celsius. The worms were harvested and washed with M9 and total RNAs were extracted from the whole bodies by using RNeasy Plus Mini (QIAGEN). Poly(A)+ RNAs were reverse transcribed with PrimeScript II (Takara) and oligo dT primer. sams-3 and sams-4 cDNA fragments were amplified first with primer pairs 5-GTTTCAACCTTACAACCAAA-3 and 5-CGGACGAGAACYTGRTAGTC-3, and 5-GACGCTTTCAAACATCCATCA-3 and 5 CGGACGAGAACYTGRTAGTC-3', respectively, by using PrimeStar GXL (Takara). The sams-3 and sams-4 cDNA fragments were further amplified with 5-TAATACGACTCACTATAGGGAGAATCTCGGACGCTGTTCTTGA-3 and 5-TTTTTTTTTTTTTTTTTTTTGGGGGGCGGACGAGAACCTGGTAGTC-3, and 5-TAATACGACTCACTATAGGGAGAATCTCGGACGCTGTTCTTGA-3 and 5-TTTTTTTTTTTTTTTTTTTTCCCCCCCGGACGAGAACCTGGTAGTC-3, respectively, to attach T7 promoter and A20 tail by using PrimeStar GXL (Takara). The cDNA fragments were utilized as templates for in vitro transcription with T7 RNA polymerase (Takara) and the RNAs were purified with RNeasy Plus Mini (QIAGEN). The RNAs were subjected to poly(A) tailing with E-PAP (Ambion). The sams-3 and sams-4 RNAs were then pooled and subjected direct RNA sequencing with Nanopore MinION platform.
description
L1
http://www.ebi.ac.uk/efo/EFO_0001084
dev stage
smg-2 (yb979) I
genotype
Caenorhabditis elegans
http://purl.obolibrary.org/obo/NCBITaxon_6239
organism
Nonsense-mediated mRNA decay (NMD)-deficient
phenotype
sams3_sams4
sample name
KH1668
strain
whole
http://purl.obolibrary.org/obo/NCIT_C25326
tissue
C.elegans_KH1668_L1_RNA_oligo(dT)_sams-3_sams-4_in-vitro-transcription_polyA
title