Academic journal article Iranian Journal of Public Health

Design of Novel Internal Positive Control (Ipc) for Molecular Detection of Infectious Bronchitis Virus (Ibv) through Real Time Rt Pcr

Academic journal article Iranian Journal of Public Health

Design of Novel Internal Positive Control (Ipc) for Molecular Detection of Infectious Bronchitis Virus (Ibv) through Real Time Rt Pcr

Article excerpt

Background: The aim of the present survey is to design and construct a plasmid containing recombinant 5'UTR and 3'UTR genes as positive control and internal positive control.

Methods: We constructed an IPC that can be amplified by the same primer pair as wild-type target RNA and the differ- ent site for probe. IPC was obtained by insertion of an exog- enous DNA fragment into reference strain target by recom- binant PCR and finally the production was colonized in pTZ57RT.Afterwards, the plasmid constructs were trans- formed into E. coli TOP10F ?host strain. Screening the desired recombinant clone was carried out using colony PCR

Results: Sequencing confirmed the presence of the desire insert (IPC sequence) in recombinant plasmid. Consequently, the IPC fragment was longer than the target gene while both ends had similar attachments to the same primer pair. The probe site is different in positive control and IPC. …

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