The Selection About Yak Hair Color Candidate Genes And MC1R Gene Functional Verification

The experiment taken Datong yak and Tianzhu white yak as objective study with different hair color.Taken fluorescence quantitative PCR,paraffin section,B16 cell model method.Used Real-time fluorescent quantitative PCR technology determi nated Agouti,MCIR,MITF and TYR gene relative expression volume in yak skin organization with different hair color;Taken toluidine blue and HE dyeing observ ated the melanin particles distribution in skin organization with different hair color;B16 cell as biological model,controlled MCIR gene expression by shRNA,lately determinated melanin particles content and Agouti,MCIR,MITF and TYR gene expression volume.Results were as follows:(1)Real-time fluorescent quantitative PCR detection got Agouti gene mRNA expression volume in black hair and white hair was not significant difference;The black hair MCIR gene mRNA expression volume was White hair 2.4 times,very significant;MITF gene mRNA expression volume in white hair was high,and compared with black hair was significant difference;TYR gene mRNA expres sion volume in black hair than white hair was higher,significant difference.Guesse d black hair and MCIR,TYR high expression levels were closely related;white co at was related with MITF expression.(2)Toluidine blue staining than HE was easier to see the distribution of melanin granules and skin layers,but not so good for melanocytic staining;HE staining was good for staining melanocytic,you can see the distribution of melanin cell size and shap e,the cell structure,cytoplasmic and nuclear.Melanin cell in the epidermal basal cell an d between its top and bottom,and also distributed around the hair follicles.HE stained Datong yak found hair follicle attached to the hair near the papilla the melanin content than other parts around hair follicle was higher.Irregular skin melanocytes,which generally has a tendency to form air bubbles,larger nuclei,mature melanocytes with inconspicuously dendritic.White hair no melanin granules around the hair follicles,but the small amount of melanin granules found in the basal layer of the epidermis.Black hair epidermal basal andaround hair follicles found large amounts of melanins.White coat around the hair follicle melanocytes melan in cause white hair can not be generated.(3)The B16 cell as a organism model,by shRNA gene silented MCIR gene in cell.Observated the melanin synthesis in the late time.After the shRNA plasmid transfected cells did not change,activity was not affected.The experiment was divide d in 3 groups:test group with specificity shRNA for objective gene.test group with noneeffective shRNA for objective gene and blank control.Transfected specifi c shRNA,B16 significantly reduced synthesis of melanin granules,and control group and control group were very significant.From protein level,the blank control and negative control respectively were test control 7.56 times and 6.83 times,the 87%suppression ratio.The MC1R gene expression,the blank control and negative control respectively were test control 11.21 times and 10.19 times.The test control was lower than the others control,very significant,above 91%suppression ratio.In the Tianzhu white yak hair follicle distributed melanophore and a little melanin grains;MC1R in melanoma cell expression played a vital role in the synthesis of melanin granules.