The Research On The Grape PC And Its ANR Gene Cloning

Proanthocyanidin(PC),or condensed tannins,belongs to flavonoids.It is a type of substance with antioxidation,Anti-tumor and a series of biological activities.As one of the world's most popular 10 medicinal plants,PC has been widely used in medicine,health products and cosmetics.Grape is rich in proanthocyanidin.The proanthocyanidin content of the 31 varieties of grape skin and grape seed were studied.The results showed that there was remarkable difference in the PC.content among different varieties.Grape seed PC contents varied between a maximum of 21.27%and a minimum of 3.83%;For proanthocyanidin content in grape skin,the highest is 8.09%and the lowest only 0.09%.Two grape varieties(NO.4 and NO.21)with obvious difference in PC content were selected to extract the grape seed PC and grape skin PC,for the study of the extract's purity and polymerization degree.Then anti-oxidation tests were conducted in vivo. The results showed:The PC from grape seed is better than from grape skin in antioxidant effect;The antioxidant effect of grape seed PC in vivo is the obvious; Grape Skin Proanthocyanidins also have antioxidant effects,but it is not significant; The two varieties of grape PC have no significant difference in vivo antioxidant effect.In this study,we used the "BR" grape variety for materials to clone the open reading frame of anthocyanidin reductase gene(ANR)through homology cloning method,and obtained a 1017 bp open reading frame.Alignment results showed that the similarities between the target sequence with those of Viti vinifera ANR and Camellia sinensisANR were 99%and 84%respectively.The recombinant pET-28a-ANR was constructed with the open reading frame inserted into pET-28a.The plasmid was expressed in E.coli BL21 and SDS-PAGE was used to detect the molecular weight of fusion protein.The results indicated that the fusion protein had expected molecular weight.