绵羊磷脂酶C zeta基因对绵羊卵母细胞孤雌激活的初步研究
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摘要
旨在应用绵羊磷脂酶C zeta(PLCζ)基因对绵羊卵母细胞进行激活,探究是否可以作为一种新的卵母细胞激活物质。本研究构建了pEGFP-N1-PLCζ重组质粒,并将质粒注入绵羊MⅡ期卵母细胞,观察及统计PLCζ对卵母细胞的激活效果和胚胎发育的影响。结果表明,pEGFP-N1-PLCζ重组质粒可在卵母细胞中表达,并可自发激活卵母细胞孤雌发育。
Sheep phospholipase C zeta(PLCζ)was used to activate sheep oocytes,to explore whether PLCζcan be used as a new kind of oocyte activator.A pEGFP-N1-PLCζrecombinant plasmid were constructed and then injected into MⅡ sheep oocytes.The effect of PLCζon activation of oocytes and embryo development were observed and counted.The results showed that pEGFP-N1-PLCζrecombinant plasmid could express PLCζin oocytes,and could spontaneously activate parthenogenetic development of sheep oocytes.
Sheep phospholipase C zeta(PLCζ)was used to activate sheep oocytes,to explore whether PLCζcan be used as a new kind of oocyte activator.A pEGFP-N1-PLCζrecombinant plasmid were constructed and then injected into MⅡ sheep oocytes.The effect of PLCζon activation of oocytes and embryo development were observed and counted.The results showed that pEGFP-N1-PLCζrecombinant plasmid could express PLCζin oocytes,and could spontaneously activate parthenogenetic development of sheep oocytes.
引文
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[21]赛务加甫,张立,彭新荣,等.绵羊精子提取物对卵母细胞的孤雌激活作用[J].西北农林科技大学学报,2006,34(7):9-12.SAI W J,ZHANG L,PENG X R,et al.A pplication of sheep sperm extract cytosolic injection in oocytes parthenogenetic activation[J].Journal of Northwest A&F University(Natural Science Edition),2006,34(7):9-12.(in Chinese)
[22]PHILLIPS S,YU Y,ROSSBACH A,et al.Divergent effect of mammalian PLC-zeta in generating Ca2+oscillations in somatic cells versus eggs[J].Biochem J,2011,438(3):545-553.
[2]姜园园,张宝修,孙婧陶,等.褪黑素对小鼠卵母细胞氧化应激及孤雌发育的影响[J].畜牧兽医学报,2013,44(7):1037-1042.JIANG Y Y,ZHANG B X,SUN J T,et al.The Effect of melaonin on the oxidative stress of oocyte and parthenogentic development in mouse[J].Acta Veterinaria et Zootechnica Sinica,2013,44(7):1037-1042.(in Chinese)
[3]刘强,贺志锐,王银龙,等.中国美利奴绵羊磷脂酶C zeta基因真核表达载体的构建及其表达研究[J].畜牧兽医学报,2014 45(4):541-546.LIU Q,HE Z R,WANG Y L,et al.Construction eukaryotic expression vector and its expression of PLCζgene from Chinese Merino sheep[J].Acta Veterinaria et Zootechnica Sinica,2014 45(4):541-546.(in Chinese)
[4]MICHELL R H,ALLAN D.Inositol cyclis phosphate as a product of phosphatidylinositol breakdown by phospholipase C(Bacillus cereus)[J].FEBS Letters,1975,53(3):302.
[5]SWANN K A.cytosolic sperm factor stimulates repetitive calcium increases and mimics fertilization in hamster eggs[J].Development,1990,110(4):1295-1302.
[6]STRICKER S A.Intracellular injections of a soluble sperm factor trigger calcium oscillations and meiotic maturation in unfertilized oocytes of a marine worm[J].Dev Biol,1997,186(2):185-201.
[7]SAUNDERS C M,LARMAN M G,PARRINGTON J,et al.PLCzeta:a sperm-specific trigger of Ca2+oscillations in eggs and embryo development[J].Sci Signal,2002,129(15):3533.
[8]SCHWARZER N,NOST R,SEYBOLD J,et al.Two distinct phospholipases C of Listeria monocytogenes induce ceramide generation,nuclear factor-κB activation,and E-selectin expression in human endothelial cells[J].J Immunol,1998,161(6):3010-3018.
[9]KATAN M.Families of phosphoinositide-specific phospholipase C:structure and function[J].BBA-Mol Cell Biol L,1998,1436(1):5-17.
[10]HEINZ D W,ESSEN L O,WILLIAMS R L,et al.Structural and mechanistic comparison of prokaryotic and eukaryotic phosphoinositide-specific phospholipases C[J].J Mol Biol,1998,275(4):635-650.
[11]IWASAKI Y,NISHIKAWA S,TSUREDA M,et al.Detection of phospholipase D on solid materials[J].Analyt Biochem,2004,329(1):157-159.
[12]杨燕燕,青格勒,达来,等.Ghrelin对绵羊早期体外胚胎发育的影响[J].畜牧兽医学报,2014,45(6):926-931.YANG Y Y,QING G L,DA L,et al.Effects of Ghrelinon the development of early ovine embryos in vitro[J].Acta Veterinaria et Zootechnica Sinica,2014,45(6):926-931.(in Chinese)
[13]郑爱燕,吴曼,尚利青,等.山羊GDF9基因在卵丘卵母细胞复合体体外成熟过程中的表达[J].畜牧兽医学报,2010,41(6):671-677.ZHENG A Y,WU M,SHANG L Q,et al.Expression of growth differentiation factor 9in goat COCs during in vitro Maturation[J].Acta Veterinaria et Zootechnica Sinica,2010,41(6):671-677.(in Chinese)
[14]孙婧陶,尹多,柳海星,等.亚硒酸钠对延边奶山羊卵母细胞体外成熟及体细胞克隆重构胚发育的影响[J].畜牧兽医学报,2013,44(11):1758-1765.SUN J T,YIN D,LIU H X,et al.Effects of sodium selenite on the yanbian dairy goat oocyte in vitro maturation and the effects on the development of somatic cell nuclear transfer embryos[J].Acta Veterinaria et Zootechnica Sinica,2013,44(11):1758-1765.(in Chinese)
[15]贺志锐,肖红冉,于振兴,等.中国美利奴绵羊磷脂酶C zeta基因的克隆及序列分析[J].动物医学进展,2013,34(5):31-36.HE Z R,XIAO H R,YU Z X,et al.Cloning and sequencing analysis of PLCζgene from Chinese Merino sheep[J].Progress in Veterinary Medicine,2013,34(5):31-36.(in Chinese)
[16]HOKIN M R,HOKIN L E.Enzyme secretion and the incorporation of P32intophospholipides of pancreas slices[J].Biol Chem,1953,203(2):967-977.
[17]赛务加甫,彭新荣,李向臣,等.精子提取液对绵羊同异种核移植重构胚的激活效果[J].畜牧兽医学报,2008,39(10):1343-1348.SAI W J,PENG X R,LI X C,et al.Effects of ovine sperm extract on the development competence of homogeneous and heterogeneous embryos reconstructed by nuclear transfer[J].Acta Veterinaria et Zootechnica Sinica,2008,39(10):1343-1348.(in Chinese)
[18]LIN T P.Microinjection of mouse eggs[J].Science,1966,151(3708):333-337.
[19]武迪迪,张杰,具英花,等.p21-EGFP融合基因表达载体与卵母细胞的表达定位[J].贵阳医学院学报,2007,32(1):5-8.WU D D,ZHANG J,JU Y H,et al.Construction of p21/EGFP fusion gene expression vector and its expression in mouse oocyte[J].Journal of Guiyang Medical College,2007,32(1):5-8.(in Chinese)
[20]COWARD K,KUBOTA H,HIBBITT O,et al.Expression of a fluorescent recombinant form of sperm protein phospholipase C zeta in mouse epididymal sperm by in vivo gene transfer into the testis[J].Fertil Steril,2006,85:1281-1289.
[21]赛务加甫,张立,彭新荣,等.绵羊精子提取物对卵母细胞的孤雌激活作用[J].西北农林科技大学学报,2006,34(7):9-12.SAI W J,ZHANG L,PENG X R,et al.A pplication of sheep sperm extract cytosolic injection in oocytes parthenogenetic activation[J].Journal of Northwest A&F University(Natural Science Edition),2006,34(7):9-12.(in Chinese)
[22]PHILLIPS S,YU Y,ROSSBACH A,et al.Divergent effect of mammalian PLC-zeta in generating Ca2+oscillations in somatic cells versus eggs[J].Biochem J,2011,438(3):545-553.