Detection of exogenous gene doping of IGF-I by a real-time quantitative PCR assay.

Abstract:

:Gene doping can be easily concealed since its product is similar to endogenous protein, making its effective detection very challenging. In this study, we selected insulin-like growth factor I (IGF-I) exogenous gene for gene doping detection. First, the synthetic IGF-I gene was subcloned to recombinant adeno-associated virus (rAAV) plasmid to produce recombinant rAAV2/IGF-I-GFP vectors. Second, in an animal model, rAAV2/IGF-I-GFP vectors were injected into the thigh muscle tissue of mice, and then muscle and blood specimens were sampled at different time points for total DNA isolation. Finally, real-time quantitative PCR was employed to detect the exogenous gene doping of IGF-I. In view of the characteristics of endogenous IGF-I gene sequences, a TaqMan probe was designed at the junction of exons 2 and 3 of IGF-I gene to distinguish it from the exogenous IGF-I gene. In addition, an internal reference control plasmid and its probe were used in PCR to rule out false-positive results through comparison of their threshold cycle (Ct) values. Thus, an accurate exogenous IGF-I gene detection approach was developed in this study.

authors

Zhang JJ,Xu JF,Shen YW,Ma SJ,Zhang TT,Meng QL,Lan WJ,Zhang C,Liu XM

doi

10.1002/bab.1518

subject

Has Abstract

pub_date

2017-07-01 00:00:00

pages

549-554

issue

4

eissn

0885-4513

issn

1470-8744

journal_volume

64

pub_type

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