Isolation and cloning of flanking regions (ndhB–rps7) of plastidial vector from lettuce

Abstract

33356003 Abstract . One of the advantages of chloroplast engineering is the entrance of targeted gene in to a specific region of the chloroplast genome. Due to advantage of this method than nuclear genetic engineering, it is one of the safest ways for the production of recombinant biopharmaceuticals, vaccines etc ., in the plants. This is done due to the presence of homologous sequences from chloroplast genome of host plant on both sides of the gene cassette and the occurrence of homologous recombination. In this research, cloning of ndhB–rps7 fragment was considered from lettuce chloroplast genome as flanking regions. The designing of specific primers for amplification of these regions was done by Primer 3 and Oligo Analyzer softwares. After total genomic DNA extraction, PCR amplification of fragment was done and cloning of this fragment was carried out by T/A procedure in to p TG19–T vector. The recombinant plasmids were sent to Genefanavaran co for sequencing and determining the restriction sites in the cloned fragment. The validation of ndhB–rps7 fragment was confirmed with the lettuce 1380 bp through the ‘BLAST’ software. This work is the first step in the production of biomaterials in