Relations On Cross-combination Of β-and α-polypeptides Of Light Harvesting Complexes II(LHII) Between Rhodobacter Sphaeroides And Rhodovulum Sulfidophilum

Rhodobacter sphaeroides and Rhodovulum sulidophilum belong to different generas of photosynthetic bacteria.β- andα-polypeptides of light harvesting complexes II(LHII) in Rhodobacter sphaeroides and Rhodovulum sulidophilum are encoded respectively by two group of homologous genes, pucB,pucA and pucsB,pucsA. The two group of genes were connected crosswise to form two new permutations, pucB-pucsA and pucsB-pucA, then cloned into the expression vector controlled by puc promoter coming from Rhodobacter sphaeroides,respectively, to yield two plasmids, PRKpucApucsB and PRKpucsApucB, which were introduced into LHI-, LHII- and RC- mutation Rhodobacter sphaeroides(DD13). The result shows that both of the two transconjugant strains can synthesize LHII complexes and construct into membrane system subsequently. The main results are as follows:①The gene fragment pucsBA was amplified by a couple of primers, then, it was cloned into pMD18-T to construct a recombinant plasmid pMD18-T-pucsBA, which was checked by sequencing.②The plasmids pMD18-T-SBA and pRKpucApuc2B*HHT were digested with SacI and XbaI, We got the gene fragment pucsB and the vector fragment pRKpucA, then, they were ligated with T4-DNA ligase, we get the expression vector-pRKpucsBpucA; the plasmids pMD18-T-SBA and pRKpucBpuc2A-1 were digested with Xbal and BamHI, we acquired the gene fragment pucsA and the vector fragment pRKpucB, with the similar method we got the expression vector-pRKpucBpucsA.③Using E. coli S17-1 as the donor, the two expression vectors were introduced into LHI-, LHII- and RC- mutation Rhodobacter sphaeroides(DD13) by conjugation, through resistance blotting, we acquired two transformants- Rb. sphaeroides DD13/pRKpucsBpucA and Rb. sphaeroides DD13/pPKpucBpucsA。④By RT- PCR, we found both of the two hybrid genes were expressed effectively.⑤Through spectral analysis, we can decide that the hybrid light-harvesting protein can combine with the light-harvesting pigment, and become the pigment-protein complexes⑥All the results above can demonstrate that the light-harvesting proteins in two kinds of photosynthetic bacteria which have different phenotypes have versatility to some extent.