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组织特异RNAi转基因小鼠模型的构建

DOI: 10.1360/052011-100, PP. 289-295

Keywords: 组织特异,RNAi,转基因小鼠模型

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Abstract:

RNAi是一种行之有效的基因沉默的新方法,被广泛地应用于基因功能的研究、疾病的治疗以及新型疫苗的研制等领域.本研究通过原核显微注射干扰载体的方法制备转基因小鼠.选用皮肤组织特异表达的人源角蛋白14(K14)基因启动子(2000bp)作为表达载体启动子,成功地驱动融合表达载体EGFP-shRNA进行干扰片段前体的转录,进而生成成熟的干扰片段,靶向小鼠BMP4基因使其发生沉默.所得到的转基因小鼠及其杂交后代经PCR和Southern杂交鉴定,结果表明外源基因准确无误地整合到小鼠基因组.Northern杂交结果证明,小干扰RNA在皮肤组织中有较高水平的表达,在肺和肠组织中有较低水平的表达.研究结果表明,利用PolⅡ型(K14)启动子驱动shRNA融合转录本的表达,在特定组织高表达siRNA,从而达到抑制特定组织目的基因表达的技术路线是可行的.同时为利用K14启动子进行毛囊相关基因干扰研究积累了基础数据,为制备组织特异抑制基因表达的转基因大家畜提供了一个参考方法.

References

[1]  21 Vivi K, Makoto M, Kazunari T, et al. Control of siRNA expression using the Cre-loxP recombination system. Nucleic Acids Res, 2004, 32:e66??
[2]  22 李福兵, 杜晓兰, 余英, 等. 骨形成蛋白4 条件性RNA 干扰小鼠的建立. 遗传, 2008, 30: 341-346
[3]  25 叶梅霞, 刘军梅, 李昊, 等. amiRNAi-实现高效稳定的特异基因沉默新方法. 中国生物工程杂志, 2010, 30: 118-125
[4]  26 袁圆阳, 骆启聪, 李超, 等. Pygo2 转基因小鼠模型的建立及表形的初步分析. 厦门大学学报(自然科学版), 2010, 49: 266-271
[5]  27 任红艳, 张兴举, 杨述林, 等. c-Myc 转基因小鼠构建的改良方法. 农业生物技术学报, 2008, 16: 37-40
[6]  23 Andrea V, Alexander M, Christopher P D, et al. Cre-lox-regulated conditional RNA interference from transgenes. Proc Natl Acad Sci USA,2004, 101: 10380-10385??
[7]  24 Frank S, Guang H, Richard J R, et al. A lentiviral microRNA-based system for single-copy polymerase II-regulated RNA interference inmammalian cells. Proc Natl Acad Sci USA, 2005, 102: 13212-13217??
[8]  1 Fire A, Xu S, Montgomery M K, et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature,1998, 391: 906-811
[9]  2 曹明媚, 任浩, 赵平, 等. 小干扰RNA 对庚型肝炎病毒基因在人Huh-7 细胞中表达折抑制作用. 中国科学C 辑: 生命科学, 2004, 34:343-349
[10]  3 Sui G C, Soohoo C, Affar E B, et al. A DNA vector-based RNAi technology to suppress gene expression in mammalian cells. Proc Natl Acad Sci USA, 2002, 99: 5515-5520??
[11]  4 王丰, 马家烈, 李惠明, 等. 靶向E2F1 的shRNA 抑制眼内视网膜细胞瘤生长. 生物化学与生物物理进展, 2009, 36: 587-591
[12]  5 娄颜坤, 罗娟, 代蓉, 等. 小鼠肌肉生长抑制素基因短发夹RNA 促进MyoD 在C2C12 细胞中表达. 生物化学与生物物理进展, 2010,37: 451-459
[13]  6 Boden D, Pusch O, Lee F, et al. Promoter choice affects the potency of HIV-1 specific RNA interference. Nucleic Acids Res. 2003, 31:5033-5038
[14]  7 Tiscornia G, Tergaonkar V, Galimi F, et al. CRE recombinase-inducible RNA interference mediated by lentiviral vectors. Proc Natl AcadSci USA, 2004, 101: 7347-7351??
[15]  8 Ventura A, Meissner A, Dillon C P, et al. Cre-lox-regulated conditional RNA interference from transgenes. Proc Natl Acad Sci USA, 2004,101: 10380-10385??
[16]  9 Hutvágner G, Zamore P D. A microRNA in a multiple-turnover RNAi enzyme complex. Science, 2002, 297: 2056-2060??
[17]  10 Zhou H, Xia X G, Xu Z. An RNA polymerase II construct synthesizes short-hairpin RNA with a quantitative indicator and mediates highlyefficient RNAi. Nucleic Acids Res, 2005, 33: e62??
[18]  11 Rao M K, Pham J, Imam J S, et al. Tissue-specific RNAi reveals that WT1 expression in nurse cells controls germ cell survival andspermatogenesis. Genes Dev, 2006, 20: 147-152??
[19]  12 Ling X, Li F Z. Silencing of antiapoptotic survivin gene by multiple approaches of RNA interference technology. Biotechniques, 2004, 36:450-458
[20]  13 Yuan J, Wang X B, Zhang Y, et al. shRNA transcribed by RNA Pol II promoter induce RNA interference in mammalian cell. Mol Biol Rep,2006, 33: 43-49??
[21]  14 Sharov A A, Weiner L, Sharova T Y, et al. Noggin overexpression inhibits eyelid opening by altering epidermal apoptosis anddifferentiation. EMBO J, 2003, 22: 2992-3003??
[22]  15 Botchkarev A V, Sharov A A, Funa K, et al. Mdodulation of BMP signaling by noggin is required for induction of the secondary(nontylotrich) hair follicle. J Invest Dermatol, 2002, 118: 3-10??
[23]  16 Andl T, Ahn K, Kairo A, et al Epithelial Bmpr1a regulates differentiation and proliferation in postnatal hair follicles and is essential fortooth development. Development, 2004, 131: 2257-2268
[24]  17 Vassar R, Rosenberg M, Ross S, et al. Tissue-specific and differentiation-specific expression of a human K14 keratin gene in transgenicmice. Proc Natl Acad Sci USA, 1989, 86: 1563-1567??
[25]  18 Seyed J H, Alireza Z, Razieh J, et al. A study of the expression of functional human coagulation factor Ⅸ in keratinocytes using a nonviralvector regulated by K14 promoter. Appl Biochem Biotech, 2010, 162: 1599-1611??
[26]  19 Purva S, Chen C, Sonali P G, et al. Loss of integrin α9β1 results in defects in proliferation, causing poor re-epithelialization duringcutaneous wound healing. J Invest Dermatol, 2009, 129: 217-228??
[27]  20 Wang X M, Sandra Z, Kenneth P, et al. Transgenic studies with a keratin promoter-driven growth hormone transgene: prospects for genetherapy. Proc Natl Acad Sci USA, 1997, 94: 219-226??

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