摘要
体细胞核移植的成功说明了高度分化的细胞仍具有支持发育的全能性,卵母细胞胞质中存在着将分化体细胞重编程至多能胚胎细胞状态的全部因子。但是,体细胞核移植的成功率却非常低,而且克隆动物存在不同程度的发育缺陷。目前普遍认为,体细胞核移植的低成功率以及发育异常和供体细胞核的不完全表观遗传重编程有关。DNA甲基化和组蛋白乙酰化是哺乳动物主要的表观遗传修饰形式,是调节基因组功能的重要手段,在胚胎的正常发育过程中具有显著的调控作用。本文对牛体外受精(IVF)胚胎和体细胞核移植(SCNT)胚胎附植前各个发育阶段的DNA甲基化和组蛋白乙酰化状态进行分析,找出体细胞克隆胚胎在基因组甲基化和组蛋白乙酰化水平上与IVF胚胎的差异,研究表观遗传状态对克隆胚体外发育能力的影响。
1.研究卵母细胞成熟培养液中添加表皮生长因子(EGF)和胰岛素(Insulin)对其体外发育的影响,精子的不同处理方法对卵泡卵母细胞体外受精的影响以及细胞共培养系统对体外受精胚胎体外发育的影响。结果表明,成熟培养液中添加30 ng/mL EGF可以显著提高卵母细胞的成熟率和受精后的卵裂率;精子经上浮法处理后可以得到较高的卵裂率和囊胚发育率;输卵管上皮细胞共培养可以显著提高受精后的囊胚发育率,可以用于牛IVF胚胎的体外培养。
2.探讨了不同激活方法对牛核移植胚胎激活效果和体外发育的影响,以及细胞共培养体系对牛体细胞核移植胚胎体外发育的影响。结果表明,5μmol/L离子霉素联合2mmol/L 6-DMAP激活牛核移植胚胎的效果比较好,可以得到较高的卵裂率和囊胚发育率;输卵管上皮细胞饲养层共培养可以显著提高重构胚的囊胚发育率,比较适合作为体细胞核移植胚胎发育的共培养体系。
3.采用免疫荧光染色技术,对牛IVF胚胎和SCNT胚胎附植前各个发育阶段的DNA甲基化和组蛋白乙酰化状态进行检测,研究克隆胚胎在基因组甲基化和组蛋白乙酰化水平上与IVF胚胎的差异。结果表明,克隆胚胎各个发育阶段DNA甲基化水平均高于体外受精胚胎,其中2-细胞到桑椹胚阶段甲基化水平显著高于IVF胚胎,从头甲基化时间提前;克隆胚胎4-细胞到囊胚阶段乙酰化水平低于体外受精胚胎,其中8-细胞阶段乙酰化水平显著低于体外受精胚胎。与体外受精胚胎相比,牛核移植胚胎附植前各阶段存在异常的DNA甲基化和组蛋白乙酰化模式。
4.用DNA甲基化酶抑制剂(5-aza-dC)处理牛供体细胞和克隆胚胎,探讨DNA甲基化水平变化与克隆胚胎体外发育之间的关系。结果表明,低浓度的5-aza-dC(20nM,72h)处理供体细胞或者重构胚不能显著提高克隆胚胎的体外发育率,对克隆胚的表观遗传状态没有明显影响;而供体细胞和重构胚均用低浓度5-aza-dC(20nM,72h)处理可以显著提高囊胚发育率和囊胚细胞数,2-细胞阶段DNA甲基化水平降低,8-细胞阶段的DNA甲基化水平显著降低且与IVF胚胎相比差异不再显著,但组蛋白乙酰化水平没有明显改变。说明克隆胚胎DNA甲基化水平影响其体外发育,降低2-细胞和8-细胞阶段的DNA甲基化水平可以提高克隆胚胎的体外发育能力。
5.用组蛋白去乙酰化酶抑制剂(TSA)处理牛供体细胞和克隆胚胎,探讨组蛋白乙酰化变化与克隆胚胎体外发育的关系。结果表明,供体细胞经50nM TSA处理12h可以显著提高克隆胚胎的囊胚发育率,而且2-细胞阶段的组蛋白乙酰化水平升高;TSA处理重构胚不能显著提高克隆胚胎的体外发育率,对重构胚的表观遗传状态没有显著影响;供体细胞和重构胚均用低浓度TSA(50nM,12h)处理可以显著提高克隆胚胎的囊胚发育率,8-细胞阶段的组蛋白乙酰化水平显著增加且与IVF胚胎相比差异不再显著,但TSA处理对DNA甲基化水平没有明显影响。说明克隆胚胎组蛋白乙酰化水平影响其体外发育,增加2-细胞和8-细胞阶段组蛋白乙酰化水平可以提高克隆胚胎的体外发育能力。
6.联合应用5-aza-dC和TSA处理牛供体细胞和重构胚,研究两种试剂联合处理对重构胚发育的影响,以及对重构胚甲基化水平和乙酰化水平的影响。结果表明,5-aza-dC(20nM, 72h)联合TSA(50nM, 12h)处理供体细胞和重构胚可以更明显的提高克隆胚胎的体外发育能力,联合处理不但显著降低了2-细胞和8-细胞阶段DNA甲基化水平,而且显著增加了组蛋白的乙酰化水平,从而使克隆胚附植前各发育阶段的表观遗传状态更接近与体外受精胚胎。说明异常的DNA甲基化和组蛋白乙酰化模式不利于克隆胚的体外发育,消除这种异常可以提高克隆胚的体外发育能力。进一步的实验表明,5-aza-dC和TSA对核移植胚胎体外发育的影响没有供体细胞特异性。
Adult somatic cell nuclei can be reprogrammed into embryonic state through nuclear transfer, cytoplasm of oocyte possesses factors essential for converting differentiated somatic nuclei into pluripotent status, however, this reprogramming process is of low efficiency mainly due to incomplete somatic nucleus epigenetic remodeling. The abnormal epigenetic modifications include DNA methylation and histone acetylation that serve as monitor of chromatin structure and gene expression. In this paper, the DNA methylation and the histone acetylation status of IVF and SCNT embryos were analysed and compared, effects of epigenetic reprogramming on the development of cloned embryos were investigated.
1. In this study, effects of EGF and Insulin, different sperm treatment methods, and different somatic cell co-culture systems on bovine in vitro fertilization (IVF) were investigated. The results showed that, the matured rate and cleavage rate were significantly increased after added 30 ng/mL EGF to in vitro culture medium; the swim-up method resulted in the best cleavage rate and blastocyst rate; and the blastocyst rate was significantly increased when the fertilized oocytes were cultured with bovine oviduct epidermic cells.
2. In this study, effects of different activation methods and somatic cell co-culture systems on in vitro development of cloned bovine embryos were investigated. The results showed that, the cleavage rate and blastocyst rate were promoted after the fused couplets acticated with 5μmol/L ionomycin followed by 2mmol/L 6-DMAP; and the blastocyst rate was significantly increased when the cloned embryos were cultured with bovine oviduct epidermic cells.
3. The DNA methylation and histone acetylation status of IVF and SCNT embryos were analysed by immunodetection method, and the differences between two kinds of embryos were investigated. The results showed that, the DNA methylation level of cloned embryos were higher than that of IVF embryos, wich was significant from 2-cell to blastocyst stage, and the precocious de novo methylation was found in cloned embryos; the histone acetylation level of cloned embryos were lower than that of IVF embryos, which became significant in 8-cell stage. The results revealed that the DNA methylation and histone acetylation status were abnormal in cloned embryos.
4. The epigenetic modification status of cloned embryos were analysed after donor cells and cloned embryos were treated with 5-aza-dC, and the relationship between DNA methylation and the development of cloned embryos were investigated. The results showed that, the in vitro development potential and epigenetic modification status of cloned embryos were not changed after the donor cells or cloned embryos were treated with low concentration of 5-aza-dC (20nM, 72h); when both donor cells and cloned embryos were treated with 5-aza-dC (20nM, 72h), the blastocyst rate and blastocyst cell number were significantly increased, and the DNA methylation level of 2-cell embryos was reduced, which became significant at 8-cell stage, however, the histone acetylation level of cloned embryos was not changed. The results revealed that the DNA methylation status was important for embryo development , and reduce the DNA methylation level at 2-cell and 8-cell stages could promote the development potential of cloned embryos.
5. The epigenetic modification status of cloned embryos were analysed after donor cells and cloned embryos were treated with TSA, and the relationship between histone acetylation and development of cloned embryos were investigated. The results showed that, the blastocyst rate of cloned embryos was significantly increased after donor cells treated with 50nM TSA(12h), and histone acetylation level of 2-cell embryos was increased; the in vitro development potential and epigenetic modification status of cloned embryos were not changed after the cloned embryos were treated with TSA; when both the donor cells and cloned embryos were treated with 50nM TSA(12h), the blastocyst rate was significantly increased, and the histone acetylation level of 8-cell embryos was dramatically increased and more closer to IVF embryos, however, the DNA methylation level of cloned embryos were not changed. The results revealed that hisone acetylation status of cloned embryos was important for embryo development , and increase the hisone acetylation level at 2-cell and 8-cell stages could promote the development potential of cloned embryos.
6. In this study, the donor cells and cloned embryos were treated with TSA and 5-aza-dC, the following epigenetic status of cloned embryos were analysed and the effects of these reagents on development of cloned embryos were investigated. The results showed that, 5-aza-dC(20nM, 72h) combined with TSA(50nM, 12h) could significantly promoted the development potential of cloned embryos, the DNA methylation level of 2-cell and 8-cell embryos was significantly reduced while the histone acetylation level significantly increased, which became more closer to IVF embryos. The results revealed that the abnormal epigenetic status of cloned embryos could reduce the subsequent development ability, whereas change the abnormal status could promoted the subsequent development. Further studies demonstrated that the beneficial effects of TSA and 5-aza-dC on cloned embryos development were not donor cell specific.
引文
[1] 王光亚,段恩奎著.山羊胚胎工程[M].天则出版社(1994).
[2] 周虚,桑润滋主编.动物繁殖生物技术[M].农业出版社(2002).
[3] 陈芳.影响牛卵母细胞体外发育能力的因素[J].动物医学进展.2002,23(1):19-22.
[4] 芮荣.山羊卵巢腔前卵泡卵母细胞体外发育与成熟的研究[D].陕西杨凌:西北农业大学.1996.
[5] Thibault C. Final stage of mammalian oocytem aturation. in biggers J and schuetz A.(eds). Oogenesis [C].Baltimore: University Park Press, 1972, pp397-411.
[6] Wassarman PM, Albertini DF. The mammalian ovum [A]. In: Knobil E and Neill JD.(eds). The physiology of reproduction[C], Second Edition. New York:Raven Press, 1994, pp74-122.
[7] 郭志勤等编著.家畜胚胎工程[M].中国科技出版社(1998).
[8] Donabue R. Maturation of the mouse oocyte in vitro.1.Sequence and timing of nuclear progression[J]. J Exp Zoo. 1968, 169:237-250.
[9] 冯怀亮.哺乳动物胚胎工程[M].长春:吉林科技出版社(1994).
[10] Motlik J, Fulka J, Flechon JE. Changes in intercellular coupling between pig oocytes and cumulus cells during maturation in vivo and in vitro[J]. J Reprod Fertil, 1986, 76:31-37.
[11] Chen L, et al. Hyaluronic acid and synthesis and gap junction endocytosis are necessary for normal expansion of the cumulus mass[J]. Mol Reprod Dev, 1990, 26:236-247.
[12] Hunter AG, Moor RM. Stage-dependent effects of inhibiting ribonucleic acids and protein synthesis meiotic maturation of bovine oocytes in vitro[J]. J Dairy Sci, 1987, 70:1646-1651.
[13] Martino A, et al. Influence of the collection technique of prepubertal goat oocytes on in vitro maturation and fertilization[J]. Theriogenology, 1994, 42:859-873.
[14] Gibbons JR, Krisher RL, Carlin SK, et al. In vitro embryo production after microinjection and ovarian dynamics following transvaginal follicular oocyteaspiration[J]. Theriogenology, 1995, 43:1129-1139.
[14] 陈大元主编.受精生物学-受精机制与生殖工程[M].科学出版社(2000).
[15] Gibbons JR, Krisher RL, Carlin SK, et al. In vitro embryo production after microinjection and ovarian dynamics following transvaginal follicular oocyteaspiration[J]. Theriogenology, 1995, 43:1129-1139.
[16] Crister ES, Leibfried-Rutledge ML and Eyestone WH. Acquisition of developmental competence during maturation in vitro[J]. Theriogenology, 1986, 25:150.
[17] Gordon I, Lu KH. Production of embryos in vitro and its impact on livestock production[J]. Theriogenology, 1990, 33:77-87.
[18] Armstrong DT, et al. Gonadotropin stimulation regimens for follicular aspiration and in vitro embryo production from calf oocytes[J]. Theriogenoligy, 1994, 42:1227-1236.
[19] Dostal J. Isolation and characterization of maturation inhibiting compound in bovine follicular fluid[J]. Reprod NutrDev, 1996, 36:681-690.
[20] Pavlok A, Hahn AL and Niemann. Fertilization and developmental competence of bovine oocytes derived from different categories of antral follicles[J]. Mol Reprod DeV, 1992, 31:63-67.
[21] Mayes MA, Sirard MA. The influence of cumulus oocyte complex morphology and meiotic inhibitors on the kinetics of nuclear maturation cattle[J]. Theriogenology, 2000, 54:911-922.
[22] Otoi T, Yamamoto K, Koyama N, et al. Cryopreservation of mature bovine oocytes by vitrification in straw[J]. Cryobiology, 1998, 37(l):77-85.
[23] Gandofi F, Milanesi E, Pocar P, et al. Comparative analysis of calf and cow oocyte during in vitro maturation[J]. Mol Reprod Dev, 1998, 49(2):168-175.
[24] 钱云,师蔚群,丁家桐.哺乳动物体外受精的研究进展[J].动物科学与动物医学.2002,17(l)26-28.
[25] Lonergan P, Monaghan P, Rizes D. Effect of follicle size on bovine oocyte qualitly in vitro[J]. Mol Reprod Dev, 1994, 37:48-53.
[26] 李荣凤.牛卵母细胞皮层颗粒荧光染色法质成熟鉴定[J].畜牧兽医学报.2000,31(3):203-210.
[27] Eppig JJ. Maintance of meiotic arrest and the induction of oocyte maturation in mouse oocyte granulosa cell comp1exes deve1oped in vitro from preantal follicles[J]. Biol Reprod, 1991, 45:824-830.
[28] Sirard MA,Lambert RD. In vitro inhibition of oocyte nuclear maturntion in the bovine[J]. Biol Reprod, 1988, 39:229-274.
[29] Wininger JD, Kort Hl. Cryopreservation of immature and mature human oocyte[J]. Semin Reoprod Med, 2002, 20: 45-50.
[30] Bird JM. 哺乳动物精子活化[J].草食家畜.1993,3:26-30.
[31] 王海滨,夏国良,李美玲等.哺乳动物卵母细胞培养体系的研究进展[J].农业生物技术学报.2000,8(3):297-310.
[32] Zhang Y, Armstrong DT. Effect of follicle-stimulating hormone and ovarian steroids during in vitro meiotic maturation on fertilization of rat oocytes[J]. Gamete Res, 1989, 23:267-277.
[33] Leibfried-Rutledge ML, Crister ES, Eyestone WH, et al. Development poteniial of bovine oocytes matured in vitro or vivo[J]. Biol Reprod, 1987, 36:376-383.
[34] Saeki K, Hoshi M, Leibfried-Ruledge ML, et al. In vitro fertilition and development of bovine oocytes matured in serum-free medium[J]. Biol Reprod, 1991, 44:256-260.
[35] Fukushima M, Fukui Y. Effectes of gonadotropins and steroids on the subsequent fertilizability of extrafollicular bovine oocyts cultured in vitro[J]. Anim Reprod Sci, 1985, 9:323-332.
[36] Coskun S, Lin YC. Mechanism of action of epidermal growth factor induced porcine oocyte maturation[J]. Mol Reprod Dev, 1995, 42:311-317.
[37] Erickson RP. Recent advances in developmental genetics: growth factors and morphogens[J]. Mol Reprod Dev 1995, 41:109-125.
[38] Brucker C, Alexander NJ, et al. Transforming growth factor-alha arguments meiotic maturation of cumulus cell-enclosed mouse oocytes[J]. Mol Reprod Dev, 1991, 28:94-98.
[39] Dekel N, Sherizly I. Epidermal growth factor induces maturation of rat follicle-enclosed oocytes [J]. Endocrinology, 1985, 116:406-409.
[40] 李兵兵,卢克焕.生长因子对卵母细胞体外成熟及早期胚胎体外发育的影响[J].广西农业大学学报.1994,13(l):11-15.
[41] Robertson L , et al. Biol Reprod, 1988, 39:797-805.
[42] 张嘉保.兽医大学学报.1990,10(2):122-127.
[43] Parrish JJ, Susko-Parrish JL, Winer MA, et al. Capacitation of bovine sperm by heparin[J]. Biol Reprod, 1988, 38:1171-1180.
[44] Tajik P, Niwa K, Murase T. Effects of different protein supplements in ferti1ization medium on in vitro penetration of cumulus-intact and cumulus-free bovine oocytes matured in culture[J]. Theriogenology, 1993, 40:949-958.
[45] GotoK, Kajihara Y, Kosaka S, et al. Pregnancies after coculture of cumulus cells with bovine embryos derived from in vitro fertilization of in vitro matured follicular oocytes[J]. J Reprod Fert, 1988, 83: 753-758.
[46] Hamono S, Kuwayama M. In vitro fertilization and development of bovine oocytes recovered from the ovaries of individual donors: a comparasion between and cutting and aspiration method [J]. Theriogenology, 1993, 39:703-712.
[47] Lu KH, Gordon I, Gallagher M, et al. Pregnancy established in cattle by transfer of embryos derived from in vitro fertilization of follicular oocytes matured in vitro[J]. Vet Rec, 1987, 121:159-260.
[48] 杨增明,谭景和.哺乳动物体外受精的精子注射法[J].国外畜牧科技.1989,16(2):21-22.
[49] Flemming AD,Kuehl TJ. Effects of temperature upon capacitation of guinea pig spermatozoa[J]. J Exp Zoo, 233:405-411.
[50] Cheng WTK, Moor RM, Polge C. In vitro fertilization of pig and sheep oocytes matured in vivo and in vitro[J]. Theriogenology, 1986, 25:146.
[51] Holt WV, North RD.Thermotropic phase transitions in the plasma membrane of spermatozoa[J]. J Reprod Fertil, 1986, 78:447-457.
[52] 谭世俭,卢克焕.受精用液种类对牛卵母细胞体外受精的影响[J].中国畜牧兽医学报.1994.
[53] Ling ZL, Lu KH. Frequency of cleavage and development of in vitro bovine oocytes fertilized in different nunbers in drops with different sperm concentration[J]. Theriogenology, 1990, 33:275.
[54] Rehman N, Collins AR, Such TK, et al. Effect of sperm exposure time on in vitro fertilization and embryo development of bovine oocytes matured in vitro[J]. Theriogenology, 1994, 41: 1447-1452.
[55] 刘健,刘健民,张嘉保.哺乳动物体外受精若干问题[J].兽医大学学报.1992,12(3):307-314.
[56] 马云.牛卵泡卵母细胞体外成熟、体外受精及受精卵体外培养的研究[D].西北农林科技大学 2001届在职人员申请硕士学位论文.
[57] Benjamin G, Brackett. Analysis of factors involved in the in vitro production of bovine embryos[J]. Theriogenology, 1993, 39:43-64.
[58] Telford NA, watson AJ, Schults GA. Transition from maternal to embryonic contral in early mammalian development:a comparison of several species[J]. Mol Reproid, 1990, 26:90-100.
[59] 高建明.牛体外受精卵体外培养研究进展(上、下) [J].草食家畜.1996,3:14-19;4:21-23.
[60] Bavister BD. Culture of preimplantion embryos: fact and artifacts[J]. Hum Reprod Update, 1995, 1: 91-148.
[61] Schin SA, et al. Two-cell block to debelopment of cultured hanster embryos is catsed by phosphate and glucose[J]. Biol Reprod , 1988, 39(4): 1183-1192.
[62] Brackett BG, Iuelke KA. Analysis of factors involved in the in vitro production of bovine embryos[J]. Theriogenology, 1993, 39:43-64.
[63] Spindle AI, Pedersen RA. Hatching attachment and outgrowth of mouse blastocysts in intro: fixed nitrogen requirement[J]. J.Gxp.Zool, 1993, 186:305-318.
[64] Lane M, Gardner DK. Mouse embryo development is differentially regulated by amino acids[J]. Reprod Fertil, 1997, 109: 153-164.
[65] Miller JGO, Schultz GA. Amino acid content of preimplantation rabbit embryos and fluids of the reprodictive tract[J]. Biol Reprod, 1987, 36:125-129.
[66] Gardner DK. Changes in requirement and utlization of nutrients durling mammalian preimplantation embryo development and their significance in embryo culture[J]. Bioscience Reports, 1985,5:393-400.
[67] Steeves TE, Gardner DK. Temporal and differential effects of amino acids on bovine embryo development in culture[J]. Biol Reprod, 1999, 61:731-740.
[68] 张靖飞,李裕强,刘月凤.动物胚胎体外培养研究进展[J].动物科学与动物医学.2002,19(2):19-22.
[69] 曹旺聚.绵羊胚胎体外培养条件的研究进展[J].国外畜牧科技.1996,23(5):27-30.
[70] Takahashi Y, First NL. In vitro development of bovine one-ell embryos: influence of glucose, lactate, pyruvate, amino acid and vitamins[J]. Theriogenology, 1992, 37: 963-978.
[71] Looney CR, Lindsey BR, Gonseth CL, et al. Commercial aspects of oocytes retrieval and in vitro fertilization (IVF) for embryo production in probleam cows[J]. Theriogenology, 1994, 41: 67-72.
[72] 钱云,丁家桐,刘红林.卵母细胞成熟的形态学特征及其调控[J].国外畜牧科技.1999,26(6):26-30.
[73] Spemann H. Embryonic Development and Induction [M]. Yale University Press, New Haven, Connecticut, 1983.
[74] Briggs R, King TJ. Transplantation of living nuclei from blastula cells into enucleated frogs-eggs [J]. Proc Natl Acad Sci, USA, 1952, 38:455-463.
[75] Gurdon JB. The developmental capacity of nuclei taken from intestinal epithelium cells of feeding tadpoles [J]. J Embryol Exp Morphol, 1962, 10:622-640.
[76] Bromball JD. Nuclear transplantation in the rabbit eggs[J]. Nature, 1975, 258:719-722.
[77] Illmense K, Hoppe PC. Nuclear transplantation in Mus musculus: Developmental potenial of nuclei from preimplantation embryos[J]. Cells, 1981, 23: 9-18.
[78] McGrath J, Solter D. Nuclear transplantation in the mouse embryo by microsurgery and cell fusion[J]. Science, 1983, 220:1300-1302.
[79] Willadsen SM. Nuclear transplantation in ovine embryos [J]. Nature, 1986, 320:63-65.
[80] Prather RS, Barnes FL, Sims MM. Nuclear transplantation in the bovine embryo: assessment of donor nuclei and recipient oocyte [J]. Biol Reprod, 1987, 37:859-866.
[81] Smith LC, Wilmut I. Influence of nuclear and cytoplasmic activity on the development in vivo of sheep embryos after nuclear transplantation [J]. Biol Reprod, 1989, 40:1027-1035.
[82] Sims M, First NL. Production of calves by transfer of nuclei from cultured inner cell mass cells[J]. Proc Nat. Acad Sci, USA 1994, 91:6143-6147.
[83] Campbell K, McWhir J, Ritchie B, et al. Production of live lambs following nuclear of culturedembryonic disc cells [J]. Theriogenology, 1995, 43:181.
[84] Bondioli KR, Westhusin ME, Looney CR. Production of identical offspring by nuclear transfer [J]. Theriogenology, 1990, 33:165-174.
[85] Stice SL, Keefer CL. Multiple generational bovine embryo cloning [J]. Biol Reprod, 1993, 48:715- 719.
[86] Kwon OY, Kono T. Production of identical sextuplet mice by transferring metaphase nuclei from Four-cell embryos [J]. Proc Natl Acad Sci, USA, 1996, 93:13010-13013.
[87] Kwon OY, Kono T, Nakahara T. Production of live young by serial nuclear transfer with mitotic stages of donor nuclei in mice. J Reprod Dev, 1997, 43:25-31.
[88] Tsunoda Y, Kato Y. Not only inner cell mass cell nuclei but also trophectoderm nuclei of mouse blastocysts have a developmental totipotency [J]. J Reprod Ferti, 1998, 113:181-184.
[89] Sotomaru Y, Kato Y, Tsunoda Y. Induction of pluripotency by injection of mouse trophectoderm cell nuclei into blastocysts following transplantation into enucleated oocytes[J]. Theriogenology, 1999, 52:213-220.
[90] 李雪峰,谭丽玲,石德顺,等.电激活对完全体外化牛细胞核移植的影响[J]. 畜牧兽医学报. 1996, 27(6): 495-500.
[91] 窦忠英,雷安民,高志敏,等.猪胚胎细胞核移植的研究.中国生物工程学会第二次会议论文摘要.1997,9
[92] 赵浩斌,陈乃清,魏庆信.猪卵母细胞的电激活[J].西北农业学报.1999,8(8):15-19.
[93] 王斌,范必勤.兔核移植胚胎的克隆研究[J]. 畜牧兽医学报.1995: 26 (1): 1-6.
[94] 周琪.家兔胚胎细胞核移植与连续移植的研究[D].博士生研究论文. 1996, 10:44-52.
[95] 周琪,李子义,谭景和,等.核质比对家兔胚胎发育的影响[J].科学通报.1999:44(3):289-291.
[96] 张涌,王建辰,钱菊汾,等.山羊卵核移植的研究[J].中国农业科学.1991,24(5):1-6.
[97] 邹贤刚,李光三,王玉阁,等.山羊胚胎细胞经继代细胞核移植后其发育能力的研究[J]. 科学通 报.1995,40(3):264-267.
[98] Czolowska R, Modlinski JM, Tarkowski AK. Behavior of thymocyte nuclei in non-activated and activated mouse oocytes[J]. Cell Sci, 1984, 69:19-34.
[99] Kono T, Kwon OY, Ogawa M, et al. Development of mouse oocytes reciving embryonic nuclei and thymocytes[J]. Theriogenology, 1991, 35: 227.
[100] Wilmut I, Schnieke AE, McWhir J, et al. Viable offspring derived from fetal and adult mammalian cells[J]. Nature, 1997, 385: 810-813.
[101] Signer EN, Dubrova YE, Jeffreys AJ, et al. DNA fringer printing Dolly[J]. Nature, 1998,394:329-330
[102] Ashworth D, Bishop M, Campbell K, et al. DNA microsatellite analysis Of Dolly[J]. Nature, 1998, 394: 329.
[103] Kato Y, Tani T, Sotomaru Y, et al. Eight calves cloned from somatic cells of a single adult[J]. Science, 1998, 282: 2095-2098.
[104] Keefer CL, Keyston R, Lazaris A, et al. Production of cloned goats after nuclear transfer using adult somatic cells[J]. Biol. Reprod, 2002, 66: 199-203.
[105] Wakayama T, Perry ACF, Zuccotti M, et al. Full term development of mice from enucleated oocytes injected with cumulus cell nuclei[J]. Nature, 1998, 394: 369-374.
[106] Chesne P, Adenot PG, Viglietta C, et al. Cloned rabbits produced by nuclear transfer from adult somatic cells[J]. Nat. Biotechnol, 2002, 20: 366-369.
[107] Campbell KHS, McWhir J, Ritchie WA, et al. Sheep cloned by nuclear transfer from a cultured cell line[J]. Nature, 1996, 380: 64-66.
[108] Wells DN, Misica PM, Tervit HR. Production of cloned calves following nuclear transfer with cultured adult mural granulosa cells[J]. Biol. Reprod, 1999, 60: 996-1005.
[109] Cibelli JB, Stice SL, Golueke PJ, et al. Cloned transgenic calves produced from nonquiescent fetal fibroblasts[J]. Science, 1998, 280: 1256-1258.
[110] Wells DN, Misica PM, McMillan WH, et al. Production of cloned bovine fetuses following nuclear transfer with cells from a fetal fibroblast cell line[J]. Theriogenology, 1998, 49: 330.
[111] Wells DN, Misica PM, Forsyth JT, et al. The use of adult somatic cell nuclear transfer to preserve the last surviving cow of the Enderby Island cattle breed[J]. Theriogenology, 1999, 51: 217.
[117] 郭继彤. 成年体细胞克隆山羊研究[D]. 西北农林科技大学博士学位论文,2000.
[118] 安晓荣,苟克勉,陈永福,等.体细胞克隆法生产绵羊转基因囊胚[J].科学通报.2001,46(10): 820-824.
[119] 成勇,王玉阁,罗金平,等. 由成年转基因山羊体细胞而来的克隆山羊[J]. 生物工程学报. 2002, 18(1): 79-83.
[120] 吕自力,毕春明.用成纤维细胞生产克隆牛的研究.草食家畜[J].2003 年(增刊):152-154.
[121] Qi Zhou, et al. Generation of Fertile Cloned Rats by Regulating Oocyte Activation[J]. Science, 2003 302: 1179.
[122] 童第周,吴尚勤,叶毓芬,等.鱼类细胞核的移植.J 科学通报,1963,57-61.
[123] 童第周,叶毓芬,陆德裕,等.鱼类不同亚科间的细胞核移植.J 动物学报 1973,19(3):201.
[124] Mc Grath J, Solter D. Nucleocytoplasm id interaction in the mouse embryos J. J Embryol Exp M orphol, 1986, 97(suppl):277-289.
[125] Wolfe BA, W esthusin ME, LevanduskiMJ, et al. Preimplantation Development of embryos produced by intergeneric nuclear transplan-tation J. Theriogenology, 1990, 33(1):350.
[126] 梅棋,邹贤刚,杜森.鼠兔核质杂交胚胎早期发育的研究.J 实验生物学报,1993, 26(4): 389- 394.
[127] Dominko T, M italipova M, Haley B, et al. Bovine oocytes cytoplasm supports Development of em- bryos produced by nuclear transfer of somatic cell nuclei from various m am m alian species J. Biology of Reproduction, 1999, 60: 1496- 1502.
[128] Whire KL, Bunch TD, M italipov S, et al. Establishm ent of pregnancy after the transfer of nuclear embryos produced from the fusion of Argali(Ovis ammon) nuclei into domestic sheep (Orvis aries) enucleated oocytes J. Cloning, 1999, 1:47- 54.
[129] 陈大元,孙青原,刘冀珑,等.大熊猫供核体细胞在兔卵胞质中可去分化而支持早期重构胚发育.J 中国科学(C 辑), 1999, 29(3): 324-330.
[130] Yoon JT, Choi EJ, Han KY, et al. In vitro Development of embryos produced by nuclear transfer ofporcine somatic cell nuclei into bovine oocytes using three different culture systems J. Theriogenology, 2001, 55: 298.
[131] Lanza RP, Cibelli JB, Diaz F, et al. Cloning of an endangered species (B as gaurus) using interspecies nuclear transfer J. Cloning, 2000, 2(2):79-90.
[132] 孙国凤(摘).将人的体细胞移植到牛卵中培育出人 ES 细胞.J 生物技术通报,1999, (2): 41-42.
[133] Loi P, Ptak G, BarboniB, et al. Genetic rescue of an endangered mammal by cross-species nuclear transfer using postmortem somaticcells J. Nature Biotechnology, 2001, 19: 962- 964.
[134] Vogel G. Cloned gaur a short-lived success[J]. Science, 2001, 291(5503):409.
[135] McCrath J, Solter D. Inability of mouse blastomere transferred to enucleated zygotes to support development in vitro [J]. Science, 1984, 226: 1317-1319.
[136] Tsunoda Y, Yasui T, Shioda Y, et al. Full-term development of mouse blastomere nuclei transplanted into enucleated two-cell embryos [J]. J Exp Zool, 1987, 242(2): 147-151.
[137] Echelard Y, Destrempes MM, Koster JA. Production of recombinant human serum albumin in the milk of transgenic cows [J]. Theriogenology, 2002, 57: 779.
[138] Chen SH, Vaught TD, Monahan JA, et al. Efficient production of transgenic cloned calves using preimplantation screening [J]. Biol Reprod, 2002 (67): 1488-1492.
[139] Cheong H, Takahashi Y, Kanagawa H. Relationship between nuclear remodelling and subsequent development of mouse embryonic nuclei transferred to enucleated oocytes [J]. Mol Reprod Dev, 1994, 37: 138-145.
[140] Booth PJ, Tan SJ, Reipurth R, et al. Simplification of bovine somatic cell nuclear transfer by application of a zona-free manipulation technique [J]. Cloning and Stem Cells, 2001, 3(3): 139-150.
[141] Yang X. Embryo cloning by nuclear transfer in cattle and rabbits [J]. Embyto Trans, Newletter, 1991, 9: 10-22.
[142] Fulka JJr, Notarianni E, Passoni L, et al. Early changes in embryonic nuclei fused to chemically enucleated mouse oocytes [J]. Int J Dev Biol, 1993, 37(3): 433-439.
[143] Baguisi A, Overstrom. Induced enucleation in nuclear transfer procedures to produce cloned animal [J]. Theriogenology, 2000, 53: 209.
[144] 杨东山. 转基因克隆法制作人胰岛素原牛乳腺生物反应器的研究[D]. 呼和浩特: 内蒙古大学博士学位论文, 2005.
[145] Wang MK, Liu JL, Li GP, et al. Sucrose pretreatment for enucleation: an efficient and non-damage method for removing the spindle of the mouse MII oocyte [J]. Mol Reprod Dev, 2001, 58(4): 432-436.
[146] Liu L, Oldenbourg R, Trimarchi JR, et al. A reliable, noninvasive technique for spindle imaging and enucleation of mammalion oocytes [J]. Nat Biotechnol, 2000, 18: 223-224.
[147] Yang CH, Yanagimachi R, Yanagimachi H, et al. Morphology and fertilizability of zona-free hamster eggs separated into halves and quarters by centrifugation [J]. Biol Reprod, 1989, 41: 741-752.
[148] Brendan G, Tatham, Kim J, et al. Electrofusion parameters for Nuclear Transfer predicated using Isofufsion Contours Produced with Bovine Embtyonic Cells [J]. Mol Repro Dev, 1996, 443: 306-312.
[149] Bordignon V, Smith LC. Telophase enucleation:an improved method to prepare recipient cytoplasts for use in bovine nuclear transfer [J]. Mol Reprod Dev, 1998, 49: 29-36.
[150] Smith LC. Membrane and intracellular effects of ultraviolet irradiation with Hoechst33342 on bovine aecondary oocytes matured in vitro [J]. J Reprod Fertil, 1993, 99: 39-44.
[151] Morgan A, Jacob R. Ionomycin enhances Ca2+ influx by stimulating store regulated cation entry and not by a direct action at the plasma membrane [J]. Biochem J, 1994, 300: 665-672.
[152] Chen DY, Jiang MX, Zhao ZJ, et al. Cloning of Asian yellow goat (C. hircus) by somatic cell nuclear transfer: telophase enucleation combined with whole cell intracytoplasmic injection [J]. Mol Reprod Dew, 2007, 74(1): 28-34.
[153] Campell KHS, Lolp, Otaegli PJ, et al. Cell cycle co-ordination in embryo cloning by nuclear transfer[J]. Reviews of Reproduction, 1996a, 1(1):40-46.
[154] Kues WA, Carnwath JW, Niemann H, et al. Serum deprivation induced DNA fragmentation of porcine fetal fibroblasts[J]. Theriogenology, 2000, 53:228 (abstr).
[155] Campell KHS. Nuclear equivalence nuclear transfer and the cell cycle[J]. Cloning, 1999, 1(1):3-15.
[156] Ono Y, Shimozawa N, Ito M, et al. Cloned mice from fetal fibroblast cells arrested at metaphase by a serial nuclear transfer[J]. Biol Reprod, 2001a:64:44-50.
[157] Ono Y, Shimozawa N, Muguruma K, et al. Production of cloned mice from embryonic stem cells arrested at metaphase[J]. Reproduction, 2001b, 122:731-736.
[158] Liu L, Dai Y, Moor RM. Nuclear transfer in sheep embryos:the efect of cell-cycle coordination between nucleus and cytoplasm and the use of in vitro matured oocytes[J]. Mol Reprod Dev, 1997, 47:255-264.
[159] Zhou Q, Jouneau A, Borcbard V, et al. Developmental potential of mouse embryos reconstructed from metaphase embryonic stem cell nuclei [J]. Biol Reprod, 2001, 65: 412-419.
[160] Rohl JM, Prater R, Barnes F, et al. Nuclear transplantation in bovine embryos [J]. J Anim Sci, 1987, 64: 642-647.
[161] GoTo K, Tanaka M, Ookutsu S, et al. Production of a nuclear transfer calf by the intracytoplasmic injection of donor cells [J]. J Reprod Dev, 1997, 43(3): 257-260.
[162] Sugimura T, Ushijima T. Genetic and epigenetic alterations in carcinogenesis[J]. Mutat Res, 2000, 462(2-3):235-246.
[163] Rickard LF, White KI. Electrofusion-induced Ca2+ flux and its effect on murine oocyte activation[J]. Mol Reprod Dev, 1992, 31:152-159.
[164] Smith LC, Wilmut I, Hunter RHF. Influence of cell cycle stage at nuclear transplantation on the development in vitro of mouse embryos[J]. J Reprod Fert, 1988, 84:619-624.
[165] Bodo S, Dinnyes A, Baranyai B, et al. Comparison of different treatments for parthenogenetic activation of bovine oocytes matured in vitro[J]. Acta Vet Hung, 1998, 46: 493-500.
[166] Machaty Z, Prather RS. Strategies for activating nuclear transfer oocytes[J]. Reprod Fertil Dev, 1998, 10:599-613.
[167] Zakhartchenko V, Alberio R, Stojkovic, et al. Adult cloning in cattle:potential of nuclei frompermanent cell line and from primary cultures[J]. Mol Reprod Dev, 1999, 54:264-272.
[168] Mitalipov SM, White KL, Farrar VR, et al. Development of nuclear transfer and parthenogenetic rabbit embryos activated with inositol 1,4,5-trisphosphate[J]. Bio Reprod, 1999, 60(4):821-827.
[169] Loi P, Ledda S, Fulka J, et al. Development of parthenogenetic and cloned ovine embryos: effect of activation protocols[J]. Biol Reprod, 1998, 58(5):1177-1187.
[170] Yoo JG, Lee HJ, Choe SY, et al. The use of cdc2 kinase inhibitor as a potent activator in reconstituted bovine embryos with primordial germ cells[J]. Theriogenology, 2001, 55:297.
[171] Wang WH, Machaty Z, Ruddock N, et al. Activation of porcine oocytes with calcium ionophore: effects of extracellular calcium[J]. Mol Reprod Dev, 1999, 53:99-107.
[172] Knott JG, Kasinathan P, Wu H, et al. Effect of porcine sperm factor on intracellular calcium and activation of bovine oocytes and development of nuclear transfer embryos[J]. Theriogenology,2001, 55:455.
[173] McGinnis LK, Youngs CR. In vitro development of ovine embryos in CZB medium. Theriodenology, 1992, 37: 559-569.
[174] Pinyopummintr T, Bavister BD. Effect of gaseous atmosphere on in vitro maturation and in vitro fertilization of bovine oocytes. Theriogenology, 1994, 41: 276.
[175] Carolan C, Lonergan P, van Langendonckt A, et al. Factors affecting bovine embryo development in synthetic oviduct fluid following oocyte maturation and fertilization in vitro. Theriogenology, 1995, 43: 1115-1128.
[176] Gardner Dk. Changes in requirements and utilization of nutrients during mammalian preimplantation embryo development and their significance in embryo culture. Theriogenology, 1998a, 49(1): 83-102.
[177] Gardner DK, Rodriguez-Martinez H, Lane M. Fetal development after transferring is increased by replacing with the glycosaminoglycan hyaluronan for mouse embryo culture and transfer. Hum Reprod, 1999, 14: 2575-2580.
[178] Ando H, Kobayashi M, Todas, et al. Establishment of a ciliated epithelial cell line from human fallopian tube. Hum Reprod, 2000, 15(7):597.
[179] Bavister BD. Role of oviductal secretions in embryonic growth in vivo and in vitro. Theriogenology, 1988, 29: 143-154.
[180] Ben-Chetrit A, Jurisicova A, Casper RF. Co-culture with ovarian cancer cell enhances human blastocyst formation in vitro. Fertil Steril, 1996, 65(3): 664-666.
[181]Campbell KHS, Ritchic WA, Wilmut I. Disappearance of maturation promoting factor and the vitro matured bovine oocyte[J]. Theriogenology, 1993, 39:199-205.
[182] 汪堃仁,薛绍白,柳惠图.细胞生物学[M]北京:北京师范大学出版社,1997.
[183] Fatemi M, Hermann A, Pradhan S, et al. The activity of the murine DNA methyltransferase Dnmt1 is controlled by interaction of the catalytic domain with the N-terminal part of the enzyme leadingtoan allosteric activation of the enzyme after binding to methylated DNA [J]. J Mol Biol, 2001, 309(5): 1189-1199.
[184] Wolffe AP, Jones PL, Wade PA. DNA demethylation [J]. Proc Natl Acod Sci USA, 1999, 96:5894-5896.
[185] Lie, Bestor TH, Jaenisch R. Targeted mutation of the DNA methyltransferase gene results in embryonic lethality [J]. Cell, 1992, 69: 915-926.
[186] Okano M, Bell DW, Haber DA, et al. DNA methyltransferases Dnmt3a and Dnmt3b are essential for denovo methylation and mammalian development [J]. Cell, 1999, 99: 247-257.
[187] Herman JG, Baylin SB. Gene silencing in cancer in assortatton with promoter hypermethylatian. N Engl J Med, 2003, 349: 2042-2054.
[188] Farrell WE, Simpson DJ, Frost SJ, et al. Methylation mechnisims in pituitary tumorigenesis [J]. Endocr Relat Cancer, 1999, 6 (4): 437-447.
[189] Feng YQ, Desprat R, Fu H, et al. DNA methylation supports intrinsic epigenetic memory in mammalian cells [J]. PLOS Genet, 2006, 2: 0461-0470.
[190] Beaujean N, Taylor J, Gardner J, et al. Effect of limited DNA methylation reprogramming in the normal sheep embryo on somatic cell nuclear transfer [J]. Biol Reprod, 2004, 71(1): 185-193.
[191] Davie JR. Covalent modifications of histories: expression from chromatin templates[J]. Curr Opin Genet Dev, 1998, 8:173-178.
[192] Finnin MS, Donigian JR, Cohen A, et al. Structures of a histone deacetylase homologue bound to the TSA and SAHA inhibitors[J]. Nature, 1999, 401:188-193.
[193] Richon VM, Emiliani S, Verdin E, et al. A class of hybrid polar inducers of transformed cell diferentiation inhibits histone deacetylases[J]. Proc Natl Acad Sci USA, 1998, 95:3003-3007.
[194] Santos F, Zakhartchenko, Stojkovic M, et al. Epigenetic marking correlates with developmental potential in cloned bovine preimplantation embryos[J]. Curr Biol, 2003, 13: 1116-1121.
[195] Enright BP, Jeong BS, Yang X, et al. Epigenetic characteristics of bovine donor cells for nuclear transfer: levels of histone acetylation[J]. Biol Reprod, 2003, 69:1525-1530.
[196] Enright BP, Kubota C, Yang X, et al. Epigenetic characteristics and development of embryos cloned from donor cells treated by trichostatin A or 5-aza-2'-deoxycytidine[J]. Biol Reprod, 2003, 69:896-901.
[197] Shi W, Hoeflich A, Flaswinkel H, et al. Induction of a senescent-like phenotype does not confer the ability of bovine immortal cells to support the development of nuclear transfer embryos[J].Biel Reprod, 2003, 69:301-309.
[198] Eggan K, Akutsu H, Honchedlinger K, et al. X chromosome inactivation in cloned mouse embryos[J]. Science, 2000, 290:1578-1581.
[199] Wakayama T, Shinkai Y Tanemura KLL, Niida H, et al. Cloning of ice to six generations[J]. Nature, 2000, 407:318-319.
[200] Lanza R P, Cibelli L B,Blackwell L, et al. Extension of cell: life-span and telmere length in animals cloned from senescent somatic cells[J]. Science, 2000, 288:665-669.
[201] Epstein CJ, Smith S, Travis B, et al. Both X chromosomes function before visible X-chromosome inactivation in female mouse embryos[J]. Nature, 1978, 274(5670):500-503.
[202] Marahrens Y, Panning B, Dausman J, et al. Xist-deficient mice are defective in dosage compensationbut not spermatogenesis[J]. Genes Dev, 1997, 11(2):156-166.
[203] Chao W, Huynh KD, Spencer RJ, et al. CTCF, a candidate trans-acting factor for X-inactivation choice[J]. Science, 2002, 295(5553):345-347.
[204] Wrenzycki C, Herrmann D, Carnwath JW, et al. Alterations in the relative abundance of gene transcripts in preimplantation bovine embryos cultured in medium supplemented with either serum or PVA[J]. Mol Reprod Dev, 1999, 53:8-18.
[205] Xue F, Tian XC, Du F, et al. Aberrant paterns of X chromosome inactivation in bovine clones[J]. Nat Genet, 2002, 31(2):216-220.
[206] Lewis IM, Peura TT, Trounson AO, et al. Pregnancy and live birth achieved from vitrified multigenerational cattle nuclear transfer embryos[J]. Theriogenology, 1998, 49: 323.
[207] Peura TT, Lewis IM, Trounson AO. The effect of recipient oocyte volume on nuclear transfer in cattle[J]. Mol Reprod Dev, 1998, 50 (2): 185-191.
[208] 安晓荣,苟克勉,朱士恩,等.卵丘细胞核移植技术生产克隆牛犊[J].中国科学(C 辑),2002, 32(1):69-76.
[209] Hochi S, Kato M, Ito K, et al. Nuclear transfer in cattle:effect of linoleic acid-albumin on 4freezing sensitivity of enucleated oocytes [J]. J Vet Med Sci, 2000, 62(10):1111-1113.
[210] Kuhholzer B, Hawley RJ, LaiL, et al. Clonal lines of transgenic fibroblast cells derived from the same fetus result in different development when used for nuclear transfer in pigs [J]. Biol Reprod, 2001, 64(6):1695-1698.
[211] Yang X, Kubota C, Suzuki H, et al. Control of oocyte maturation in cow-biological factors[J]. Theirogenology, 1998, 49(2):471-482.
[212] Adenot PG, Szolfost, Chesne P, et al. In vivo aging of oocyte sin-fluence the behavior of nuclei transferred to enucleated rabbit oocytes[J]. Mol Reprod Dev, 1997, 46(3):325-336.
[213] Keda I, Takahashi Y. Effects of maturation nalage of porcine oocytes on the induction of activation and development in vitro following somatic cell nuclear transfer[J]. J Vet Med Sci, 2001, 63(9): 1003-1008.
[214] Westhusin ME, Burghardt RC, Ruglia JN, et al. Potential for cloning dogs[J]. Reprod Fertil Suppl, 2001, 57:287-293.
[215] Enright BP, Taneja M, Schreiber D, et al. Reproductive characteristics of cloned heifers from adult somatic cells[J]. Biol Reprod, 2002, 66(2):291-296.
[216] Kato Y, Tani T, Tsunoda Y. Cloning of calves from various somatic cell trpes of male and female adult, newborn and fetal cows[J]. J Reprod Fertil, 2000, 120: 231-237.
[217] Tetsuya Tani, Yoko Kato, Yu kioTsunoda. Direct exposure of chromosomes to nonactivated ovum cytoplasm is effective for bovine somatic cell nucleus reprogramming [J]. Biol Reprod, 2001, 64: 324-330.
[218] Wakayama T, Yanagimachi R. Effect of cytokinesis inhibitors, DMSO and the timing of oocyte activation on mouse cloning using cumulus cell muclei[J]. Reproduction, 2001, 122(1):49-60.
[219] Watson AJ, De Sousa P, Caveney A, et al. Impact of bovine oocyte maturation media on oocytetranscript levels, blastocyst development, cell number, and apoptosis[J]. Biol Reprod, 2000, 62:355-364.
[220] Niemann H, Wrenzycki C. Alterations of expression of developmentally important genes in preimplantation bovine embryos by in vitro culture conditions:implications for subsequent development[J]. Theriogenology, 2000, 53:21-34.
[221] Stojanov T, O’Neill C. Ontogeny of expression of a receptor for platelet-activating factor in mouse preimplantation embryos and the effects of fertilization and culture in vitro on its expression[J]. Biol Reprod, 1999, 60:674-682.
[222] Parrish JJ, Krogenaes A, Susko-Parrish JL, et al. Effect of bovine sperm separation by either swim-up or percoll method on success of in vitro fertilization and early embryonic development[J]. Theriogenology, 1995, 44:859-869.
[223] 徐照学.牛卵泡卵母细胞体外受精及体外受精胚冷冻保存的研究[D].西北农业大学 1995 届攻读博士学位研究生学位论文.
[224] Patk KW, Iga K, Niwa K. Exposure of bovine oocytes to EGF during maturation allows them to develop to blast oocysts in a chemically-defined medium[J]. Theriogenology, 1997, 48:1127-1135.
[225] Guler A, Poulin N, Mermillod P, et al. Effect of growth factors, EGF and IGF-I and estradiol on in vitro maturation of sheep oocytes[J]. Theriogenology, 2000, 54 (2) :209-218.
[226] 李向臣,丛日华,安志兴,等.EGF 和 FBS 对牛孤雌胚胎发育的影响[J].西北农林科技大学学:自然科学版,2004,32(4):51-54.
[227] Lonergan P, Carolan C, Van Langendonekt A, et al. Role of epidermal growth factor in bovine oocyte maturation and preimplantation embryo development in vitro[J]. Biol Reprod, 1996, 54:1420.
[228] Lorenzo PL, Illera MJ, Illera JC , et al. Enhancement of cumulus expansion and nuclear maturation during bovine oocyte maturation in vitro by the addition of epidermal growth factor and insulin-like growth factor I[J]. J Reprod Fertil, 1994, 101 (3) :697-701.
[229] Ilera M J, Lorenzo P, Illera J C, et al. Epidermal growth factor increase the bovine oocyte maturation in vitro. 12th International Congress on Animal Reproduction, 1992, 339-341.
[230] 季尚娟,刘铁铮.生长因子对体外受精—胚胎移植的影响[J]当代畜牧,2003,9:5-6.
[231] Zhang L. Blackwood EG, Denniston RS, et al. The effect of insulin on maturation and Development of in vitro fertilized bovine oocytes. Theriogenology, 1991b.35:301.
[232] Zhang L, Denniston RS, Godke RA. The effect of insulin on the Development of in vitro fertilized pocine oocytes. J. Anim. Sci, 1991(supple 1).
[233] Matsui M, Takahashi Y, Hishinuma M, et al. Insulin and insulin-like growth factor-I I(IGF-I) stimulate the Development of bovine embryos fertilized in vitro. J. Vet. Med. Sci, 1995,57(6):1109-1111.
[234] 文国艺.牛体外受精四种方法的比较[J].广西畜牧兽医.2000,16(2):5-7.
[235] Kane MT. The role of nutrients, peptide growth factors and co-culture cells in development of preimplantation embryos in vitro[J]. Theriogenology, 1992, 38: 297-313.
[236] 冯怀亮,李彦舫,刘建民.牛卵母细胞体外受精及体外发育的研究[J].生物技术.1993,3(3):18-21.
[237] 王秀阁,谭美容.牛体外受精技术研究[J].湖北畜牧兽医.1995,3:1-3.
[238] 罗光彬,韩雪花,金花子.体外培养条件对延边黄牛受精卵体外发育的影响[J].中国兽医学报.2004,24(2):192-194.
[239] Kang YK, Koo DB, Park JS, et al. Typical demethylation events in cloned pig embryos. J Biological Chemistry, 2001, 276(43): 39980-39984.
[240] San Buissho A, Threlfall WR. The effects of estrous cow serum on the in vitro maturation and fertilization of bovine follicular oocytes [J]. Theriogenology, 1989, 31:300-309.
[241] Critser ES, First NL. Use of a fluorescent stain for visualization of nuclear material in living oocytes and early embryos [J]. Stain Technology, 1986, 61:1-5.
[242] Liu, FJ, Zhang, Y, Zheng YM, et al. Optimization of electrofusion protocols for somatic cell nuclear transfer. Small Rumin. Res. 2007, 73, 246-251.
[243] Alberio R, Kubelka M, Zakhartchenko V, et al. Activation of bovine oocytes by specific inhibition of cyclin-dependent kinases[J]. Mol Re-prod Dev, 2000, 55(4):422-432.
[249] Prather RS, Eichen PA, Nicks DK, et al. Artificial activation of porcine oocytes matured in vitro. Molecular Reproduction Development [J], 1991, 28: 405-409.
[250] Dinneyes A, Dal Y, Jiang S, et al. High development rates of vitrified bovine oocytes following parthenogenetic activation, in vitro fertilization, and somatic cell nuclear transfer[J]. Bio Reprod, 2000, 63: 513-518.
[251] Szollosi D, Czolowska R, Szollosi MS, et al. Remodelling of mouse thymocyte nuclei depends on the time of their transfer into activated, homologous ocytes[J]. J cell Sci, 1988, 91:603-613.
[252] Rafael A, Fissore, James M. Intracellular Ca2+ response of rabbit oocytes to electrical stimulation[J]. Mol Reprod Dev, 1992, 32:9-16.
[253] Satoh T, Kobayashi, Yamashita S, et al. Tissue inhibitor of metalloproteinase(TIMP-1) production by granulose and oviduct cells enhances in vitro development of bovine embryo[J]. Biol Reprod, 1994, 50:835-844.
[254] Bird AP, Wolffe AP. Methylation-induced repression-belts, braces and chromatin. Cell, 1999, 99: 451-454.
[255] Baguisi A, Behboodi E, Melican DT. Production of goats by somtic cell nuclear transfer. Nat Biotechnol, 1999, 17:456-461.
[256] Polejaeva IA, Chen Sh, Vaught TD, et al. Cloned pigs produced by nuclear transfer from adult somatic cells. Nature, 2000, 407(6800):86-90.
[257] Onishi A, Iwamoto M, Akita T, et al. Pig cloning by microinjection of fetal fibroblast nuclei. Science 2000, 289(5482):1188-1190.
[258] Daniels R, Hall V, Trounson AO. Analysis of gene transcription in bovine nuclear transfer embryos reconstructed with granulose cell nuclei. Biol Reprod, 2006, 3(4):1034-1040.
[259] Humpherys D, Eggan K, Akutsu H, et al. Abnormal gene expression in cloned mice derived from embryonic stem cell and cumulus cell nuclei. Proc Natl Acad Sci USA, 2002, 99:12889-12894.
[260] Li E. Chromatin modification and epigenetic reprogramming in mammalian development. Nat Rev Genet, 2002, 3(9):662-673.
[261] Kang YK, Koo DB, Park JS, et al. Aberrant methylation of donor genome in cloned bovine embryos. Nat Genet, 2001, 28:173-177.
[262] Ohgane J, Wakayama T, Kogo Y, et al. DNA methylation variation in cloned mice. Genesis 2001, 30:45-50.
[263] Shi W, Dirim F, Wolf E, et al. Methylation reprogramming and chromosomal aneyploidy in in vivo fertilized and cloned rabbit preimplantation embrys. Biol Reprod, 2004, 71(1):340-347.
[264] Mann M, Chung Y, Nolen L, et al. Disruption of imprinted gene methylation and expression in cloned preimplatation stage mouse embryos. Biol Reprod, 2003, 69(3):902-914.
[265] Wee G, Koo DB, Song BS, et al. Inheritable Histone H4 Acetylation of Somatic Chromatins in cloned Embryos[J]. Journal of Biological Chemical, 2006, 281:6048-6057.
[266] Zhang L, Wang SH, Fan BL, et al. Global histone H4 acerylation of IGF1 and GH genes in lungs of somatic cel cloned calves. Asian_aust. J.Anim.Sci, 2006, 19(8):1090.
[267] Slack A, Cervoni N, Pinard M, et al. Feedback regulation of DNA methyltransferase gene expression by methylation [J]. Eur J Biochem, 1999Aug, 264(1):191-199.
[268] Jones PA, Takai D. The role of DNA methylation in mammalian epigenetics. Science, 2001,293: 1068-1070.
[269] Rideout III WM, Eggan K, Jaenisch R. Nuclear cloning and epigenetic reprogramming of the genome. Science, 2001, 293:1093-1098.
[270] Lee J, Jnoue K, Ono R, et al. Erasing genomic imprinting memory in mouse clone embryos produced from day 11.5 primordial germ cells. Development, 2002, 129:1807-1817.
[271] Santos F, Dean W. Epigenetic reprogramming during early development in mammals[J]. Reproduction, 2004, 147:643-651.
[272] Dean W, Bowden L, Aitchison A, et al. Altered imprinted gene methylation and expression in completely ES cell-derived Mouse fetuses: association with aberrant phenotypes. Development, 1998, 125(12): 2273-2282.
[273] Dean W, Santos F, Stojkovic M, et al. Conservation of methylation reprogramming in mammalian development:aberrant reprogramming in cloned embryos. Proc Natl Acad Sci USA, 2001, 98(24): 13734-13738.
[274] Kang YK, Park JS, Koo DB, et al. Limited demethylation leaves mosaic-type methylation states in cloned bovine pre-implantation embryos. EMBO J, 2002, 21(5): 1092-1100.
[275] Bourc’his D, Le Bourhis D, Patin D, et al. Delayed and in complete reprogramming of chromosome methylation patterns in bovine cloned embryos. Curr Biol, 2001, 11(19): 1542-1546.
[276] Reik W, Dean W, Walter J. Epigenetic reprogramming in mammalian development. Science, 2001, 293(5532):1089-1093.
[277] Huang C, Sloan EA, Boerkoel CF. Chromatin remodeling and human disease[J]. Curr Opin Genet Dev,2003, 13(3):246-252.
[278] Richon VM, Emiliani S, Verdin E, et al. A class of hybrid polar inducers of transformed cell differentiation inhibits histone deacetylases. Proc Natl Acad Sci USA, 1998, 95: 3003-3007.
[279] Yang FK, Hao R, Kessler B, et al. Rabbit somatic cell cloning:effects of donor cell type, histone acetylation status and chimeric embryo complementation[J]. Reproduction, 2007, 133:219-230.
[280] Enright BP, Sung LY, Chang CC, et al. Methylation and acetylation characteristics of cloned bovine embryos from donor cells treated with 5-aza-2’-deoxycytidine. Biol Reprod, 2005, 72:944-8.
[281] Suteevun T, Parnpai R, Smith SL, et al. Epigenetic characteristics of cloned and in vitro-fertilized swamp buffalo (Bubalus bubalis) embryos. J Anim Sci, 2006, 84:2065-71.
[282] Jaenisch R, Bird A. Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nat Genet, 2003, 33(Supplement):245-54.
[283] 沈羽非.染色质与表观遗传调控.高等教育出版社.2005.
[284] Hu JF, Nguyen PH, Pham NV, et al. Modulation of IGF2 genomic imprinting in mice induced by 5-azacytidine, an inhibitor of DNA methylation. Mol Endocrinol, 1997, 11(13):1891-1898.
[285] Pedone PV, Pikaart MJ, Cerrato F, et al. Role of histone acetylation and DNA methylation in the maintenance of the imprinted expression of the H19 and IGF2 genes. FEBS Lett, 1999, 458(1):45-50.
[286] El Kharroubi A, Piras G, Stewart CL. DNA demethylation reactivates a subset of imprinted genes in uniparental mouse embryonic fibroblasts. J Biol Chem, 2001, 276(12):8674-8680.
[287]Wijermans P, Lubbert M, Verhoef G, et al. Low-dose 5-aza-2′-deoxycytidine, a DNA hypomethylating, agent, for the treatment of high-risk myelodysplastic syndrome: a multicenter phaseⅡ study in elderly patients[J]. J Clin Oncol, 2002, 18(5):956-962.
[288] 方晓明,孙立峰,彭佳萍,等.5-氮-2′-脱氧胞苷对RKO结肠癌细胞株P16/CDKN2基因去甲基化的转录调节作用[J].中华医学杂志,2003,83(23) :2077-2082.
[289] 佟红艳,林茂芳.5′-杂氮-2′-脱氧胞苷对高危组骨髓增生异常综合征细胞作用的体外研究[J].中国实验血液学杂志,2004 ,12(4):467-471.
[290] Daskalakis M, Nguyen TT, Nguyen C, et al. Demethylation of a hypermethylated P15/ IN K4B gene in patients with myelodysplastic syndrome by 5-aza-2′-deoxycytidine (decitabine) treatment[J]. Blood, 2002, 100(8):2957-2964.
[291] Lubbert M, Wi J ermans P, Kunzmann R, et al. Cytogenetic responses in highrisk myelodysplastic syndrome following lowdose treatment with the DNA met hylation inhibitor 5-aza-2-deoxycytidine[J]. Br J Haematol, 2001, 114(2):349-357.
[292] Robertson KD, Wolffe AP. DNA methylation in health and disease. Nat Rev Genet, 2000, 1(1):11?19.
[293] Bestor TH. The DNA methyltransferases of mammals. Hum Mol Genet, 2000, 9(16): 2395-2402.
[294] Chung YG, Ratnam S, Chillet JR. Abnormal regulation of DNA methyltransferase expression in cloned mouse embryos[J]. Biol Reprod, 2003, 69:146-153.
[295] Berger SL. Histone modifications in transcriptional regulation. Curr Opin Genet, 2002, 12:142-148
[296] Strahi BD, Allis CD. The language of covalent histone modification. Nature, 2000, 403:41-45.
[297] Grunstein M. Histone acetylation in chromatin structure and transcription[J]. Nature, 1997, 389(6649):349-352
[298] Eberharter A, Becker PB. Histone acetylation: a switch between repressive and perimissive chromatin: Second in revies series on chromatin dynamics[J]. EMBO Rep, 2002, 3:224-229.
[299] Yoshida M, Hendzel M, Akita M, et al. Potent and specific inhibition of mammalian histone deacetylase both in vivo and in vitro by trichostatin A. Biol Chem. 1990, 265:17174-17179.
[300] Kishigami S, Mizutani E, Ohta H, et al. Significant improvement of mouse cloning technique by treatment with trichostatin A after somatic nuclear transfer. Biochem Biophys Res Commun, 2006, 340:183-9.
[301] Rybouchkin A, Kato Y, Tsunoda Y. Role of histone acetylation in reprogramming of somatic nuclei following nuclear transfer. Biol Reprod, 2006, 74:1083-9.
[302] Annunziato AT, Hansen JC. Role of histone acetylation in the assembly and modulation of chromatin structures. Gene Expr, 2000, 9(1-2):37-61.
[303] Zhao Q, Cumming H, Cerruti L, et al. Site-specific acetylation of the fetal globin activator NF-E4 prevents its ybiquitination and regulates its interaction with the histone deacetylase, HDAC1. J Biol Chem, 2004, 279(40):41477-41486.
[304] Yang XJ, Seto E. Collaborative spirit of histone deacerylases in regulating chromatin structure and gene expression. Curr Opin Genet Dev, 2003, 13(2):143-153.
[305] Marks PA, Richon VM, Rifkind RA. Histone Deacetylase Inhibitors:inducers of diffrentiation or apoptosis of transformed cells. J Natl Cancer Inst, 2000, 92:1210-1216.
[306] GilchristS, GilbertN, PerryP, et al. Nuclear organization of centromeric domains is not perturbed by inhibition of histone deacetylases. Chromosome Res, 2004, 12(5):505-516.
[307] Taddei A, Roche D, Sibarita JB, et al. Duplication and maintenance of heterochromatin domains. J Cell Biol, 1999, 147(6): 1153-1166.
[308] de Ruijter AJM, van Gennip AH, Caron HN, et al. Histone deacetylases (HDACs): characterization of the classical HDAC family. Biochem J, 2003, 370:737-749.
[309] Drummond DC, Noble CO, Kirpotin DB, et al. Clinical development of histone deacetylase inhibitors as anticancer agents. Annu Rev Pharmacol Toxicol, 2005, 45:495-528.
[310] Brush MH, Guardiola A, Connor JH, et al. Deactylase inhibitors disrupt cellular complexes containing protein phosphatases and deacetylases [J]. J Biol Chem, 2004, 279(9):7685-7691.
[311] Yoshida M, Horinouchi S, Beppu T. Trichostatin A and trapoxin: novel chemical probes for the role of histone acetylation in chromatin structure and function. Bioessays, 1995, 17(5): 423-430.
[312] Zhang Y, Li J, Villemoes K, et al. An epigenetic modifier results in improved in vitro blastocyst production after somatic cell nuclear transfer. Cloning Stem Cells, 2007, 9: 357-363.
[313] Shi LH, Miao YL, Ouyang YC, et al. Trichostatin A (TSA) improves the development of rabbit-rabbit intraspecies cloned embryos, but not rabbit-human interspecies cloned embryos. Dev Dyn, 2008, 237(3): 640-648.
[314] Cervoni N, Szyf M. Demethylase activity is directed by histone acetylation[J]. Biol Chem, 2001, 276: 40778-40787.
[315] German TM, Robertson KD. Chromatin remodeling,histone modifications and DNA methylation-how does it fall together[J]. J.Cell Biochem, 2002, 87:117-125.
[316] Nan X, Ng H-H, Johnson CA, et al. Transcriptional repression by the methyl-Cp G-binding protein MeCP2 involves a histone deacetylase complex. Nature, 1998, 393:386-389.
[317] Johes PL, Veenstra GJC, Wade PA, et al. Methylated DNA and MeCP2 recruithistone deacetylase to repress transcription. Nat Genetics, 1998, 19:187-191.
[318] Fuks F, Burgers WA, Brehm A, et al. DNA methyltransferase Dnmt1 associates with histone deacetylase activity. Nat Genet, 2000, 24:88-91.
[319] Selker EU. Trichostatin A causes selective loss of DNA methylation in Neurosporn. Proc Natl Acad Sci USA, 1998, 95:9430-9435.
[320] Shaker S, Bernstein M, Momparler LF, et al. Preclinical evaluation of antineoplastic activity of inhibitors of DNA met hylation(5-aza-2’-deoxycytidine) and histone deacetylation (trichostatin A, depsipeptide) in combination against myeloid leukemic cells[J]. Leuk Res, 2003, 27(5):437-444.
[321] Cameron EE, Bachman KE, Myohanen S, et al. Synergy of demethylation and histone deacetylase inhibition in the reexpression of genes silenced in cancer. Nature Genetics, 1999, 21:103-107.
[322] Yin H, Blanchard KL. DNA methylation represses the expression of the human erythropoietin gene by two different mechanisms[J]. Blood, 2000, 95(1):111-119.