Explore The Correlation Between KIF2A Gene And Differentially Expressed Genes And Screen The Pathogenic Gene Of BBS Family In Miao Nationality

Objective:Explore the correlation between the KIF2A gene and the cilia-related genes such as BBS1,BBS7,ANXA1,KIF3A and ?-Tublin,and study whether KIF2A gene have an effect on the cell proliferation or not,then screen the pathogenic genes of collected BBS family in Miao nationality and identified the results by Sanger sequencingMethod:KIF2A recombinant plasmid or siRNAs were transfected into 293T and 3T3-L1 cells,the total RNA and protein were extracted,and then the expression of KIF2A was detected by qRT-PCR and Western blotting.Analyze the correlation between the KIF2A gene and the cilia-related genes such as BBS1,BBS7,ANXA1 or cell proliferation.Then try to knock out KIF2A gene by Crispr-Cas9.The whole blood DNA of BBS pedigree were extracted,Exome sequencing of the proband and her mother were done by the Beijing Genomics Institute?BGI?.The specific location of pathogenic genes in BBS family were screened by bioinformatics analysis.Whole blood DNA of 300 Dai nationality,300 Hani nationality,300 Yao nationality and 76 Miao nationality were extracted,then BBS7 fragment was amplificated by PCR,and sequenced by Sanger sequencing.Result:KIF2A gene and siRNAs were successfully transfected into the 293T or 3T3-L1 cell.The correlation among KIF2A and BBS1,BBS7,ANXA1,KIF3A,?-Tublin and other cilia-related genes was detected.In 293T cells,when KIF2A gene overexpressed,the expression of BBS7 gene was lower than control group,and the expression of KIF3A gene was higher than control group.In 3T3-L1 cells,when KIF2A gene overexpressed,the expression of ANXA1 gene was higher than control group.The KIF2A knockout vector was constructed by Crispr-Cas9,and the knockout effect of KIF2A gene will be further verified.In the cell proliferation experiment,we found that overexpression of KIF2A gene promoted the proliferation of 293T and 3T3-L1 cells in a certain extent.A homozygous c.389₃90delAC?RefSeq NM₁76824.2?germline mutation in BBS7 gene was detected in this Miao pedigree,no mutation was detected in other 976 controls.Conclusion:In this study,the overexpression of KIF2A gene in 293T cells may affect the expression of BBS7 and KIF3A genes.The overexpression of KIF2A gene in 3T3-L1 cells may affect the expression of ANXA1 gene.However,it still need to be further studied the interaction among these genes.A homozygous deletion mutation in BBS7 was reported in a Chinese Miao pedigree,which expand the spectrum of human disease-causing gene with mutation of BBS7.