Sample 82
Homo sapiens
Protocols: miRNA-Seq libraries are performed from at least 1ug of extracted total RNA with a RIN greater than 7. Before starting, total RNAs are purified with miRNeasy kit wich allows the selection of the small RNA fraction less than 100b. first, a 3' adenylated DNA adaptor is ligated to the enriched sample in the absence of ATP preventing the self-ligation of miRNAs.Then a 5' RNA adaptor is ligated in the presence of ATP at the other end of the miRNAs.The RT primer complementary of the 3' adaptor is added at this stage with which it will form a duplex thereby reducing the ligation between adaptors.A reverse transcription is therefore perfomed from the RT primer and finally these captured miRNAs are amplified by PCR with primers complementary to the 3' and 5' adaptors.During this PCR a specific barcode is incorporated allowing individualisation of each library.Each PCR is loaded on the Fragment Analyzer (AATI) for a precise quantification of each miRNA peak of interest.Based on these results a equimolar pool of about ten of different samples are performed.Finally the pooled PCR product is loaded on PAGE in order to excise the band of miRNA that is extracted and purified on a Qiagen MinElute column.Finally the pooled PCR product is loaded on PAGE in order to excise the band of miRNA that is extracted and purified on a Qiagen MinElute column.
ENA-FIRST-PUBLIC
2015-02-02T17:02:27Z
ENA-LAST-UPDATE
2018-03-08T22:13:09Z
External Id
SAMEA3171472
INSDC center name
Programme Carte d'Identitite des Tumeurs
INSDC first public
2015-02-02T17:02:27Z
INSDC last update
2018-03-08T22:13:09Z
INSDC status
public
Submitter Id
E-MTAB-2833:Sample 82
age
59 year
broker name
ArrayExpress
common name
human
disease
Pheochromocytoma
individual
HS_060
overall survival delay
21 month
overall survival event
0
sample name
E-MTAB-2833:Sample 82
scientific_name
Homo sapiens
sex
male