Identifying Cry genes that determine insect resistance by multiplex polymerase chain reaction method in transgenic maize

Abstract

Aims. Multiplex PCR was developed for detection of genetic sequences that determine insect resistance (cry genes) in experimental maize samples and reference samples (containing the appropriate transformation events). Methods. The total DNA was extracted from maize living plant tissue. Isolation and purification of DNA was carried out using CTAB method. PCR was performed by [1]. Electrophoresis of each sample (12 µl) after mPCR performed in agarose gel (1,2%) during 1,5 hour at 3,5 V/cm. Results. Primers to detect genes cryIA(b), cry1F, cry1A.105, mcry3A, cry2Ab2, cry3Bb1, cry34Ab1, cry35Ab1 were designed and selected. Multiplex for simultaneous determination of few genes were worked out. mPCR amplification of sequences was carried out to determine the presence or absence of transgenes (cry). Conclusions. Multiplex PCR was developed using reference samples. Primers to detect cry genes/transgenic maize events were designed and selected. No cry genes were found out in experimental maize samples.

Key words: maize, cry genes, transformation events, multiplex PCR.