獭兔Slc7a11基因过表达对黑色素相关基因的影响及SNP检测

doi:10.16036/j.issn.1000-2650.2019.04.017

摘要/Abstract

摘要： 【目的】为了深入探究Slc7a11基因对獭兔毛色生成的影响。【方法】运用直接测序结合PCR-SSCP分子标记技术检测163只獭兔Slc7a11基因12个外显子的多态性,同时通过构建pEGFP-N1-Slc7a11过表达载体,转染RAB-9细胞,分析Slc7a11过表达对黑色素生成基因的影响。【结果】兔Slc7a11基因CDS区序列1 509 bp,经预测为碱性不稳定疏水性蛋白;多态性分析发现,Slc7a11基因在4种毛色的獭兔群体中不存在SNP位点,说明Slc7a11在獭兔群体中非常保守;进一步功能研究发现Slc7a11过表达后,黑色素生成相关基因MC1R、SLC24A5相对于阴性对照组,试验组表达量显著增加（P<0.05）。【结论】Slc7a11基因可影响黑色素生成相关基因的表达进而影响黑色素形成。

关键词: 獭兔, Slc7a11, 多态性, 过表达, 黑色素

Abstract: 【Objective】 To investigate the effect of Slc7a11 gene on hair color formation in Rex Rabbits. 【Method】 In this study, the polymorphisms of 12 exons of 163 Rex Rabbit Slc7a11 genes were detected by direct sequencing and PCR-SSCP molecular marker technology. At the sam e time, the overexpression vector of pEGFP-N1-Slc7a11 was constructed and transfected into RAB-9 cells to analyze the effect of Slc7a11 overexpression on melanogenesis gene. 【Result】 The CDS sequence of Slc7a11 gene in the rabbits was 1 509 bp, which was predicted to be an alkaline unstable hydrophobic protein. Polymorphism analysis showed that there was no SNP locus in the four coat-colored Rex Rabbit populations, indicating that Slc7a11 was very conservative in Rex Rabbit populations. Further functional studies showed that MC1R, SLC24A5 were related to melanin production after Slc7a11 overexpression compared with the negative control group. The expression level in the test group also increased significantly(P<0.05). 【Conclusion】 Slc7a11 gene can affect the expression of melanogenesis-related genes and melanin formation. The results laid a foundation for further study on the effect of Slc7a11 gene on hair color of Rex Rabbits.

Key words: Rex Rabbit, Slc7a11, polymorphism, overexpression, melanin

中图分类号:

S829.1

陈秋燃, 李佳丽, 刘明, 周娟, 陈阳, 吴信生. 獭兔Slc7a11基因过表达对黑色素相关基因的影响及SNP检测[J]. 四川农业大学学报, 2019, 37(04): 550-556.

CHEN Qiuran, LI Jiali, LIU Ming, ZHOU Juan, CHEN Yang, WU Xinsheng. Effect of Overexpression of Slc7a11 Gene in Rex Rabbit on Melanin Related Genes and SNP Detection[J]. Journal of Sichuan Agricultural University, 2019, 37(04): 550-556.

参考文献

[1] CASTLE W E.The genetics of domestic rabbits : a manual for students of mammalian genetics and an aid to rabbit breeders and fur farmers[J]. Nature, 1930, 127(3214): 847-848.
[2] 杨翠军, 葛剑, 陈赛娟,等. 彩色獭兔agouti位点基因突变及蛋白结构改变对其毛色的影响[J]. 浙江农业学报, 2015, 27(12): 2071-2077.
[3] SEARLE A G.Comparative genetics of coat colour in mammals[J]. London, UK: Logos Press,1968: 308.
[4] AIGNER B, BESENFELDER U, MATHIAS M, et al.Tyrosinase gene variants in different rabbit strains[J]. Mammalian Genome, 2000, 11(8): 700-702.
[5] FONTANESI L, TAZZOLI M, BERETTI F, et al.Mutations in the melanocortin 1 receptor(MC1R)gene are associated with coat colours in the domestic rabbit(Oryctolagus cuniculus)[J]. Animal Genetics, 2010, 37(5): 489-493.
[6] FONTANESI L, SCOTTI E, ALLAIN D, et al.A frameshift mutation in the melanophilin gene causes the dilute coat colour in rabbit (Oryctolagus cuniculus) breeds[J]. Animal Genetics, 2014, 45(2): 248-255.
[7] 朱谋, 王继文. 鸭色素合成相关基因TYR外显子1单核苷酸多态性及其与羽色性状相关性分析[J]. 四川农业大学学报, 2018, 36(2): 133-139.
[8] EWEN F KIRKNESS,VINEET BAFNA,AARON L Halpern,et al.The dog genome: survey sequencing and comparative analysis[J]. Science, 2003, 301(5641): 1898-903.
[9] COOK A L, CHEN W, THURBER A E, et al.Analysis of cultured human melanocytes based on polymorphisms within the SLC45A2/MATP, SLC24A5/NCKX5, and OCA2/P loci[J]. Journal of Investigative Dermatology, 2009, 129(2): 392-405.
[10] LIU X F, LUO J, HU X X, et al.Repression of Slc24a5 can reduce pigmentation in chicken[J]. Frontiers in Bioscience, 2011, 3(1): 158-165.
[11] 李洪涛, 何新, 周志勇,等. Slc7a11基因在不同被毛颜色哈萨克羊皮肤组织中的表达分析[J]. 遗传, 2012, 34(10): 1314-1319.
[12] 张俊珍, 董常生, 范瑞文,等. 哺乳动物毛色形成研究进展[J]. 动物医学进展, 2006, 27(s1): 65-68.
[13] 孙笑尉, 张雪莲, 李振,等. 染色质重塑因子BPTF、BRG1在不同毛色绵羊皮肤的表达与定位[J]. 畜牧兽医学报, 2017, 48(3): 446-453.
[14] 王准. Gas在绵羊耳部不同毛色皮肤中的表达与定位[D]. 太原: 山西农业大学, 2016.
[15] BARSH G, COTSARELIS G.How hair gets its pigment[J]. Cell,2007, 130(5): 779-781.
[16] ITO S, WAKAMATSU K.Chemistry of mixed melanogenesis--pivotal roles of dopaquinone[J]. Photochemistry & Photobiology, 2010, 84(3): 582-592.
[17] ITO S, WAKAMATSU K.Human hair melanins: what we have learned and have not learned from mouse coat color pigmentation[J]. Pigment Cell & Melanoma Research, 2011, 24(1): 63-74.
[18] HE X, LI H T, ZHOU Z Y, et al.Production of brown/yellow patches in the Slc7a11 transgenic sheep via testicular injection of transgene[J]. Journal of Genetics and Genomics, 2012, 39(6): 281-285.
[19] 苗鲁旭, 孙守荣, 席冬梅,等. Slc7a11基因与动物色素的形成[J]. 家畜生态学报, 2009, 30(4): 1-5.
[20] CHINTALA S, LI W, LAMOREUX M L, et al.Slc7a11 gene controls production of pheomelanin pigment and proliferation of cultured cells[J]. Proc Natl Acad Sci U S A, 2005, 102(31): 10964-10969.
[21] 何军敏, 孙军, 余雄,等. MC1R与Slc7a11基因多态性与哈萨克羊毛色相关的研究[J]. 中国畜牧杂志, 2017, 53(10): 39-42.
[22] 任玉红, 任彬, 范瑞文,等. 羊驼MC1R基因克隆及其在不同毛色个体中表达水平研究[J]. 畜牧兽医学报, 2009, 40(6): 841-845.
[23] GARCÍABORRÓN J C, SÁNCHEZLAORDEN B L, JIMÉNE ZCERVANTES C. Melanocortin-1 receptor structure and functional regulation[J]. Pigment Cell & Melanoma Research, 2010, 18(6): 393-410.
[24] LAMASON, R L.SLC24A5, a putative cation exchanger, affects pigmentation in zebrafish and humans[J]. Science, 2005, 310(5755): 1782-1786.
[25] 翁巧琴, 王进波, 王国朋,等. 特种獭兔毛色相关基因多态性的研究[J]. 中国养兔杂志, 2015(3): 4-9.
[26] 穆琳. Slc7a11在獭兔皮毛黑色素沉积中的作用机制研究[D]. 扬州: 扬州大学, 2018.