Cln8p Interaction Protein Screening, Eukaryotic Expression And Detection

Objective CLN8 is a pathogenetic gene of the neuronal ceroid lipofuscinoses(NCLs). To research the function of CLN8P, yeast two-hybrid system is employed as the approach to study the protein.All those works may provide new clues for the molecular pathogenetic mechanism. Methods. To construct the bait plasmid of CLN8 gene in yeast two hybrid system and examine whether the bait plasmid of CLN8 has self - activating effect. And then, We screen CLN8P interacted proteins from The Human Fetal Brain cDNA library .extract positive clones and analyses by bioinformatics techniques.Results The recombinant plasmid of plexA -CLN8was constructed successfully and it didn't display self - activating effect . forty-one positive clones was screened. There are several positive clones Which code ND5(NADH dehydrogenase subunit5 ,ND5). Conclusion 1 CLN8Pand ND5 maybe have interaction2 there are interaction betweenCLN8P and ND5, which is a reason of the accumulation of autofluorescent materialPartII expressing and testing CLN8P in eukaryocyteObjective This step paves the way for verifing the interactionbetween cln8 and ND5 in mammal cells.Methods The fragments of CLN8 gene was amplified by PCR. The amplifiedfragment of CLN8 was ligased with the P-Flag vector to constructrecombinant plasmid P-Flag~cln8. The recombinant plasmid was transfectedinto HeLa cell and test by Western blotResults The fragments of CLN8 gene was amplified and ligased with P-Flag,Digested with endonuclease and sequenced , the recombinant plasmid of...