WT1和FGF9基因在克隆猪隐睾中mRNA表达和DNA甲基化状态分析

doi:10.11843/j.issn.0366-6964.2014.06.009

Abstract

Abstract:

In order to find out the possible causes of the cryptorchidism in cloned pigs which produced by our research group.The RT-PCR techniques and bisulfite salt sequencing analysis(BSP) was used to analysis the expression and promoter methylation status of the promoter CpG sites of WT1 and FGF9 genes in the testis of cloned boars.The results showed that，the expression of both genes in cloned pig C1 and C2 were higher than those of control group N，expression of WT1 and FGF9 genes in cloned pig C1 was much higher (3.49，9.83 vs 1.00).Bisulfite sequencing PCR results showed that there were no significant change of WT1 gene methylation in cloned pig C1 and C2，and the methylation level of the both CpG inslands in FGF9 gene of cloned pig C1 was much higher than those in N（94.54% vs 18.18%，71.11% vs 26.67%）.The cloned pig C2 was not significant.Results indicated that the abnormal expression of WT1 gene may be one of the reasons for cryptorchidism of the cloned pigs，but the WT1 gene methylation had no relation on this abnormal expression.The change of DNA methylation reprogramming of FGF9 gene occurred in the testis of cloned pigs，which effected normal expression levels of the FGF9 gene，suggested that it may be an important factor for the occurrence of cryptorchidism in the cloned pigs.

Key words: cryptorchidism, cloning pigs, methylation, bisulfite sequencing

CLC Number:

S828.2

LI Fang-fang，DAI Jian-jun，WU Cai-feng,ZHANG Shu-shan，ZHANG De-fu，GU Xiao-long，WU Yun-long，WANG Xin. mRNA Expression and DNA Methylation Status of WT1 and FGF9 Genes in Cryptorchidism of Cloned Pigs[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2014, 45(6): 918-925.

References

［1］郭磊，李慧，韩之明.DNA 甲基化和组蛋白修饰在克隆动物发育过程中的作用［J］.遗传，2010，32(8)：762-768.
［2］KISHIGAMI S，WAKAYAMA S，HOSOI Y，et al.Somatic cell nuclear transfer：infinite reproduction of a unique diploid genome ［J］.Exp Cell Res，2008，314(9)：1945-1950.
［3］CIBELLI J B，CAMPBELL K H，SEIDEL G E，et al.The health profile of cloned animals ［J］.Nat Biotechnol，2002，20(1)：13-14.
［4］MACLAUGHLIN D T，DONAHOE P K.Sex determination and differentiation ［J］.N Engl J Med，2004，350(4)：367-378.
［5］SCHARNHORST V，VAN DER EB A J，JOCHEMSEN A G.WT1 proteins：functions in growth and differentiation ［J］.Gene，2001，273(2)：141-161.
［6］EGGERS S，SINCLAIR A.Mammalian sex determination - insights from humans and mice ［J］.Chromosome Res，2012，20(1)：215-238.
［7］COLVIN J S，GREEN R P，SCHMAHL J，et al.Male-to-female sex reversal in mice lacking fibroblast growth factor 9 ［J］.Cell，2001，104(6)：875-889.
［8］魏延昌.克隆猪印迹基因表达及其甲基化状态研究［D］.哈尔滨：东北农业大学，2010.
［9］吴志强，谢一妮，戴建军，等.H19 和 IGF2R 基因在体细胞克隆猪各组织中的甲基化状态［J］.中国兽医学报，2012，32(6)：922-926.
［10］KREMENSKOY M，KREMENSKA Y，SUZUKI M，et al.DNA methylation profiles of donor nuclei cells and tissues of cloned bovine fetuses ［J］.J Reprod Dev，2006，52(2)：259-266.
［11］LIN L，LI Q，ZHANG L，et al.Aberrant epigenetic changes and gene expression in cloned cattle dying around birth ［J］.Bmc Dev Biol，2008，8(1)：8-14.
［12］陈洁，李冬杰，张萃，等.Mash2基因在体细胞克隆牛肺脏中 DNA 甲基化状态分析［J］.Prog Biochem Biophys，2010，37(9)：960-966.
［13］BIASON-LAUBER A.Control of sex development ［J］.Best Pract Res Cl En，2010，24(2)：163-186.
［14］LIM H N，HUGHES I A，ROSS HAWKINS J.Clinical and molecular evidence for the role of androgens and WT1 in testis descent ［J］.Mol Cell Endocrinol，2001，185(1)：43-50.
［15］廖冰，林秀坤.FGF9调节性腺分化研究进展［J］.动物医学进展，2009，30(5)：80-83.
［16］SCHMAHL J，KIM Y，COLVIN J S，et al.FGF9 induces proliferation and nuclear localization of FGFR2 in Sertoli precursors during male sex determination ［J］.Development，2004，131(15)：3627-3636.

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mRNA Expression and DNA Methylation Status of WT1 and FGF9 Genes in Cryptorchidism of Cloned Pigs

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