HSP Gene Cloning And Transformation System Study Of Spirulina. Innermongoliansis

ABSTRACTSpirulina. innermongoliansis is a high quality and normal temperature adaptive variety, which was found in salt lake in Inner Mongolia. Because of its later discovering history, and its special living conditions and other related reasons, S. innermongoliansis have not be widely studied and exploited in Molecular Genetics. To improve its heat resistance can expand its breeding range and reduce the farming costs has a great value in practical production.Some researches show that heat shock protein in heat resistance of algae plays an important role. This study cloned 3 HSP family genes in S. innermongoliansis by PCR technology:HSP70,HSP90,DnaJ2 gene, and then analysis the gene sequence and its encoding protein gene sequence and its evolutionary relationship, which indicate that the target gene was cloned successfully.phycocyaninβ-gene promoter BetaP is Cloned from Inner Mongolia Spirulina, which is a strong constitutive expression promoter, the experiment proved it has a strong starting role.Using pBLUNT as the starting plasmid to construct a expression vector for three conjugative transferation:Selected constitutive promoter BetaP and added SD sequence(GGAGAG)to 5'; and adjusted the SD sequence and initiation codon ATG a distance of six bases, then connected gene DnaJ2. Triparental conjugative expression vector in E.coli were successfully expressed, indicating that the expression vector pACYC-BB was successfully constructed. Followed by a series of Inner Mongolia Spirulina transformation experiments, conjugative transfer into the three conditions were explored, and accumulated some valuable experience. Currently, transgenic research in Inner Mongolia Spirulina still in its infancy, this study can provide some theoretical basis and reference for future ratelative research.