比较牛催乳素与生长激素对外源基因表达的影响

中国生物工程杂志

2013, Vol. 33

Issue (8): 91-97

技术与方法

比较牛催乳素与生长激素对外源基因表达的影响

闫亚彬¹, 蔡勤^1,2, 蔡琳琳^1,2, 龚秀丽^1,2, 朱怡文^1,2, 管翌华^1,2, 黄英^1,2

1. 上海市儿童医院上海交通大学附属儿童医院上海医学遗传研究所上海 200040;
2. 卫生部医学胚胎分子生物学重点实验室暨上海市生殖与胚胎工程重点实验室上海 200040

Comparison of the Impact on the Transgene Expression Either by Exogenic Bovine Prolactin, or with Growth Hormone

YAN Ya-bin¹, CAI Qin^1,2, CAI Lin-lin^1,2, GONG Xiu-li^1,2, ZHU Yi-wen^1,2, GUAN Yi-hua^1,2, HUANG Ying^1,2

1. Shanghai Children’s Hospital, Shanghai Children's Hospital of Shanghai JiaoTong University, Shanghai Institution of Medical Genetics, Shanghai 200040, China;
2. Key Laboratory of Medical Embryo and Molecular Biology of the Ministry of Health and Shanghai Key Laboratory of Reproduction and Embryo Engineering, Shanghai 200040, China

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摘要： 目的: 用牛生长激素(bGH)和牛催乳素(bPRL)乳腺特异载体DNA分别转染携带人转铁蛋白(hTF)的转基因小鼠乳腺上皮细胞,比较这二种载体对外源基因(人转铁蛋白)的表达影响,为研制特异、高效表达外源蛋白转基因动物过程中载体的合理构建,提供有价值的理论和技术基础。方法:将构建β-酪蛋白启动子(P1A3)调控的bGH和bPRL乳腺特异表达载体分别转染体外培养的转基因小鼠乳腺上皮细胞;并将转染pcDNA3.1质粒和未转染细胞作为对照,收集不同代数的细胞及上清液,用RT-PCR及Elisa法分析mRNA及蛋白,分析比较二者对hTF表达水平的影响。结果:RT-PCR显示人转铁蛋白转录电泳条带明显;Elisa表明二者均能提高hTF蛋白水平的表达。结论:首次应用牛生长激素和催乳素对外源基因表达影响作用进行比较研究;β-酪蛋白启动子(P1A3)调控的bGH和bPRL均能促进在乳腺上皮细胞水平上外源蛋白表达。因此,在研制转基因动物——乳腺生物反应器中,采用bGH和bPRL促进乳汁中外源蛋白特异、高效表达的载体都将是可选择的新技术途径。

关键词： 牛生长激素; 牛催乳素; 人转铁蛋白; 乳腺生物反应器

Abstract: Objective: Transgenic murine mammary epithelial cells carrying exogenic human transferrin gene, were transfected with either bovine prolactin (bPRL)-expressing plasmid, or human growth hormone (bGH) expressing cassette driven by goat β-casein promoter. Human transferrin expression levels were then compared in order to have a useful information guiding for effectively generation of transgenic big animals. Methods: bPRL-and bGH-expressing plasmids were constructed, respectively, driven by goat-casein promoter (P1A3). Upon transfection them into transgenic murine mammary epithelial primary cells, the cells and the culture supernatants were collected with the passages. hTF mRNA and protein expression levels were analyzed and compared by RT-PCR and Elisa. Results: Both bPRL and bGH could enhance hTF expressing effectively in the transfected mammary epithelial cells evidenced by RT-PCR, the ELISA results showed that both gene products could enhance hTF transgene expression with no statistically significant difference between them. Conclusion: It was reported for the first time that both bPRL and bGH could be used for enhancing transgene expression. Therefore, they could be a good choice used for effectively expressing transgene in mammary gland bioreactor in transgenic cattles.

Key words: Bovine prolactin Bovine growth hormone Human trasferrin Mammary gland bioreactor

收稿日期: 2013-04-01 出版日期: 2013-08-25

ZTFLH:

Q786

基金资助: 转基因生物新品种培育科技重大专项资助项目(2009ZX08007-003B)

通讯作者: 黄英,E-mail:hying23@163.com E-mail: hying23@163.com

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引用本文:

闫亚彬, 蔡勤, 蔡琳琳, 龚秀丽, 朱怡文, 管翌华, 黄英. 比较牛催乳素与生长激素对外源基因表达的影响[J]. 中国生物工程杂志, 2013, 33(8): 91-97.

YAN Ya-bin, CAI Qin, CAI Lin-lin, GONG Xiu-li, ZHU Yi-wen, GUAN Yi-hua, HUANG Ying. Comparison of the Impact on the Transgene Expression Either by Exogenic Bovine Prolactin, or with Growth Hormone. China Biotechnology, 2013, 33(8): 91-97.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2013/V33/I8/91

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