Cloning and expression of lipid transfer protein-2 (LTP2) gene from Iranian rice

Lipid transfer proteins (LTPs) are a family group of proteins with low average molecular weight in plants that have numerous potential applications especially in drug delivery systems. The aim of this research was cloning of LTP coding sequence in a prokaryotic expression vector to produce a fusion protein with GST in E. coli (TOP10). As there is no intron segment in rice LTP2 gene، specific primers were implemented to introduce BamHI and XhoI restriction sites at the head and tail of amplified LTP2 coding sequence respectively، using extracted rice genome as a template. After treatment by respected restriction endonucleases، the amplified fragment was inserted into the BamHI-XhoI sites in pGEX-6p-2 vector downstream of GST to make a chimeric DNA for further production of GST-LTP fusion protein. At the next step recombinant vector was transformed into TOP10 E. coli cells for amplifying purposes. To ensure the accuracy of gene cloning، extracted recombinant vector were sent for sequencing the inserted fragment. Data revealed that cloned fragment was bona fide sequence of rice LTP2 as was reported to NCBI. Results showed that PCR amplification of GC-rich DNA sequences could be improved by Betaine، DMSO and AMS. Finally expression of LTP2 in E. coli was confirmed by SDS-PAGE technique.