研究报告

TSA促进转基因猪体细胞核移植胚胎发育和外源基因表达

展开
  • 东北农业大学生命科学学院, 哈尔滨 150030

收稿日期: 2010-10-15

  修回日期: 2010-12-21

  网络出版日期: 2011-07-25

基金资助

转基因生物新品种培育科技重大专项(编号:2008ZX08006-002, 2009ZX08006-001B)和东北农业大学“创新团队”发展计划项目资助

TSA improve transgenic porcine cloned embryo development and transgene expression

Expand
  • College of Life Science, Northeast Agricultural University of China, Harbin 150030, China

Received date: 2010-10-15

  Revised date: 2010-12-21

  Online published: 2011-07-25

摘要

不完全的表观遗传重编程是造成转基因克隆动物效率低下的主要原因, 组蛋白修饰作为表观遗传修饰的一个重要部分, 可以直接影响克隆胚胎的发育和外源基因的表达情况。TSA(Trichostatin A)作为一种组蛋白去乙酰化抑制剂, 可以改变组蛋白的乙酰化水平, 促进表观遗传重编程, 提高克隆动物的效率。同时TSA能改变染色质结构, 使转录因子易于与DNA序列结合, 促进外源基因的表达。文章确定了TSA处理转基因猪成纤维细胞和核移植胚胎的最佳条件, 分别为250 nmol/L、24 h和40 nmol/L、24 h, 通过进一步正交实验发现, TSA同时处理供体细胞和克隆胚胎可以显著的促进核移植胚胎的体外发育。此外, 无论TSA处理转基因猪成纤维细胞或核移植胚胎, 都可以提高外源基因的表达水平。

本文引用格式

孔庆然,朱江,黄波,郇延军,王峰,石永乾,刘仲凤,武美玲,刘忠华 . TSA促进转基因猪体细胞核移植胚胎发育和外源基因表达[J]. 遗传, 2011 , 33(7) : 749 -756 . DOI: 10.3724/SP.J.1005.2011.00749

Abstract

Uncompleted epigenetic reprogramming is attributed to the low efficiency of producing transgenic cloned animals. Histone modification associated with epigenetics can directly influence the embryo development and transgene expression. Trichostatin A (TSA), as an inhibitor of histone deacetylase, can change the status of histone acetylation, improve somatic cell reprogramming, and enhance cloning efficiency. TSA prevents the chromatin structure from being condensed, so that transcription factor could binds to DNA sequence easily and enhance transgene expression. Our study established the optimal TSA treatment on porcine donor cells and cloned embryos, 250 nmol/L, 24 h and 40 nmol/L, 24 h, respectively. Furthermore, we found that both the cloned embryo and the donor cell treated by TSA resulted in the highest development efficiency. Meanwhile, TSA can improve transgene expression in donor cell and cloned embryo. In summary, TSA can significantly improve porcine reconstructed embryo development and transgene expression.

参考文献

[1] Vajta G. Somatic cell nuclear transfer in its first and sec-ond decades: successes, setbacks, paradoxes and perspectives. Reprod Biomed, 2007, 15(5): 582-590.
[2] Lee GS, Kim HS, Hyun SH, Lee SH, Jeon HY, Nam DH, Jeong YW, Kim S, Kim JH, Han JY, Ahn C, Kang SK, Lee BC, Hwang WS. Production of transgenic cloned piglets from genetically transformed fetal fibroblasts selected by green fluorescent protein. Theriogenology, 2005, 63(4): 973-991.
[3] Watanabe S, Iwamoto M, Suzuki SI, Fuchimoto D, Honma D, Nagai T, Hashimoto M, Yazaki S, Sato M, Onishi A. A novel method for the production of transgenic cloned pigs: electroporation-mediated gene transfer to non-cultured cells and subsequent selection with puromycin. Biol Reprod, 2005, 72(2): 309-315.
[4] Lai LX, Kang JX, Li RF, Wang JD, Witt WT, Yong HY, Hao YH, Wax DM, Murphy CN, Rieke A, Samuel M, Linville ML, Korte SW, Evans RW, Starzl TE, Prather RS, Dai Y. Generation of cloned transgenic pigs rich in omega-3 fatty acids. Nat Biotechnol, 2006, 24(4): 435-436.
[5] Li RF, Lai LX, Wax D, Hao YH, Murphy CN, Rieke A, Samuel M, Linville ML, Korte SW, Evans RW, Turk JR, Kang JX, Witt WT, Dai YF, Prather RS. Cloned transgenic swine via in vitro production and cryopreservation. Biol Reprod, 2006, 75(2): 226-230.
[6] Vajta G, Zhang YH, Macháty Z. Somatic cell nuclear transfer in pigs: recent achievements and future possibilities. Reprod Fertil Dev, 2007, 19(2): 403-423.
[7] Pratt SL, Sherrer ES, Reeves DE, Stice SL. Factors influ-encing the commercialisation of cloning in the pork in-dustry. Soc Reprod Fertil Suppl, 2006, 62(1): 303-315.
[8] Kang YK, Koo DB, Park JS, Choi YH, Chung AS, Lee KK, Han YM. Aberrant methylation of donor genome in cloned bovine embryos. Nat Genet, 2001, 28(2): 173-177.
[9] Kang YK, Yeo S, Kim SH, Koo DB, Park JS, Wee G, Han JS, Oh KB, Lee KK, Han YM. Precise recapitulation of methylation change in early cloned embryos. Mol Reprod Dev, 2003, 66(1): 32-37.
[10] Turner BM. Histone acetylation and an epigenetic code. Bioessays, 2000, 22(9): 836-845.
[11] Rice JC, Allis CD. Histone methylation versus histone acetylation: new insights into epigenetic regulation. Curr Opin Cell Biol, 2001, 13(3): 263-273.
[12] Shogren-Knaak M, Ishii H, Sun JM, Pazin MJ, Davie JR, Peterson CL. Histone H4-K16 acetylation controls chromatin structure and protein interactions. Science, 2006, 311(5762): 844-847.
[13] Li XP, Kato Y, Tsuji Y, Tsunoda Y. The effects of trichostatin A on mRNA expression of chromatin structure-, DNA methylation-, and development-related genes in cloned mouse blastocysts. Cloning Stem Cells, 2008, 10(1): 133-142.
[14] Kishigami S, Mizutani E, Ohta H, Hikichi T, van Thuan N, Wakayama S, Bui HT, Wakayama T. Significant im-provement of mouse cloning technique by treatment with trichostatin A after somatic nuclear transfer. Biochem Biophys Res Commun, 2006, 340(1): 183-189.
[15] Iager AE, Ragina NP, Ross PJ, Beyhan Z, Cunniff K, Rodriguez RM, Cibelli JB. Trichostatin A improves his-tone acetylation in bovine somatic cell nuclear transfer early embryos. Cloning Stem Cells, 2008, 10(3): 371-380.
[16] Zhang YH, Li J, Villemoes K, Pedersen AM, Purup S, Vajta G. An epigenetic modifier results in improved in vitro blastocyst production after somatic cell nuclear transfer. Cloning Stem Cells, 2007, 9(3): 357-363.
[17] Li J, Svarcova O, Villemoes K, Kragh PM, Schmidt M, Bøgh IB, Zhang Y, Du Y, Lin L, Purup S, Xue Q, Bolund L, Yang H, Maddox-Hyttel P, Vajta G. High in vitro development after somatic cell nuclear transfer and trichostatin A treatment of reconstructed porcine embryos. Theriogenology, 2008, 70(5): 800-808.
[18] Kong QR, Wu ML, Huan YJ, Zhang L, Liu HY, Bou G, Luo YB, Mu YS, Liu ZH. Transgene expression is associated with
文章导航

/