A speedy method to detect inserted DNA fragment in cell lines transfected with retroviral vectors.

Abstract:

:Cells transfected by retroviral vectors are brought in agene of particular interest and are very useful in avariety of experiments. It is essential to testify that theDNA fragment was successfully introduced into the cellstogether with the retroviral vectors. Polymerase chainreaction is believed to be a fast and convenient method forthis purpose when using primers flanking the cloning siteof the inserted DNA. Unfortunately, a single PCR reactionoften fails to amplify the targeted fragment because of theexistence of endogenous virus DNA in cell genome. However,in this study we conducted a procedure for a single PCR,using vector-specific primers as well as a nested PCR, andsuccessfully detected the DNA fragments cloned in MFGretroviral vectors in 22 transfected cell lines. We alsoproved that real time quantitative PCR in combination withMFG-specific primer is useful to determine copy number ofthe retroviral vector in murine producer cell lines.

journal_name

Cytotechnology

journal_title

Cytotechnology

authors

Ding LH,Iimura E,Saijo K,Hamada H,Ohno T

doi

10.1023/A:1008187310534

subject

Has Abstract

pub_date

2000-11-01 00:00:00

pages

243-52

issue

3

eissn

0920-9069

issn

1573-0778

journal_volume

34

pub_type

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