Association Of Olr1Polymorphism Gene With Meat Traits And Interaction With BTA-MIR-370in Qinchuan Cattle

The oxidized low-density lipoprotein receptor1(OLR1) gene plays an important role in the degradation of the oxidized low-density lipoprotein, which causes damage to the arterial endothelium. In this study, we analyzed the association between OLR1genetic polymorphisms and meat traits in Qinchuan cattle. The results had shown that there existed a g.10497C>A mutation in OLR13’untranslated region (UTR) and the mutation site can be recognized by Pstl enzyme. Therefore, the genotypes of OLR1gene were identified by PCR-RFLP and we found three genotypes in Qinchuan cattle, including AA, AC and CC genotypes. For loin eye area and marbling depth, the individuals of CC genotype was higher than that of AA genotype (P<0.05), while there was no difference between individuals of CC and AC genotype (P>0.05). The result indicated that CC genotype had significant effect on eye muscle area and the depth of the eye muscle in Qin Chuan cattle and OLR1gene could be taken as a candidate gene affecting Qin Chuan cattle meat traits.For further investigating the mechanism of OLR1gene polymorphism affecting on meat traits, the relative expression of OLR1gene in mutant (CC genotype) and wild type (AA genotype) was detected by qPCR. The result revealed that the expression level of OLR1gene in mutant (CC genotype) individuals was significantly higher compared with that of AA genotype.Because the binding site of bta-miR-370was on the OLR1g.10497C> A mutation, so we hypothesised that this mutation in OLR13’UTR could caused binding sites of bta-miR-370absence. Thereby, to verify the interaction between the3’UTR C>A mutation of OLRl gene and bta-miR-370, and to investigate whether this C>A mutation was related with meat traits or not in Qinchuan cattle, we amplified a360-bp fragment of OLR13’UTR from AA, CC individuals respectively by PCR, inserted it into the downstream of firefly luciferase (luc+) ORF to get the luciferase reporter construct. At the same time, the bta-miR-370eukaryotic expression vector was constrcuted. Subsequently, the3’UTR reporter construct, bta-miR-370eukaryotic expression vector and pRL-TK Renilla luciferase construct were co-transfected into HEK293cells and the luciferase report gene activity was tested by using a dual-luciferase reporter assay. Results showed that the luciferase activity in AA construct reduced compared with the negative control and positive control (CC genotype) group. The result indicated that with g.10497C>A mutation, the target site of bta-miR-370was absence, while without the g.10497C>A mutation, bta-miR-370could combine with OLRl3’UTR and prompt its degradation, which may result in OLRl gene is regulated by the bta-miR-370no longer, leading to changes in protein expression levels and causing the variation of meat traits.