花青素合成调控转录基因作为直观标记的载体构建及其在玉米幼胚中的瞬时表达

生物技术通报 ›› 2014, Vol. 0 ›› Issue (4): 77-82.

花青素合成调控转录基因作为直观标记的载体构建及其在玉米幼胚中的瞬时表达

赵欣梅^1，2, 吴树彪², 王亦学², 崔贵梅², 王晓清², 杜建中², 王小丽^1，2, 尚勇进², 孙毅^1，2，3

(1.山西大学生物工程学院，太原 030006；
2.山西省农业科学院生物技术研究中心，太原 030031；
3.农业部黄土高原作物基因资源与种质创制重点实验室，太原 030031)

收稿日期:2013-10-11 出版日期:2014-04-29 发布日期:2014-04-29
作者简介:赵欣梅，女，硕士研究生，研究方向：作物遗传育种；E-mail：zxm880329@126.com
基金资助:
国家转基因生物新品种培育重大专项(2013ZX08003-001)，国家自然科学基金项目(31240081)，山西省科技攻关项目 (20110311009)，山西省财政支农项目(2011NYGX-05)

The Vector Construction of Anthocyanins as a Visual Marker Gene and Its Transient Expression in Maize Immature Embryos

Zhao Xinmei^1，2, Wu Shubiao², Wang Yixue², Cui Guimei², Wang Xiaoqing², Du Jianzhong², Wang Xiaoli^1，2, Shang Yongjin², Sun Yi^1，2，3,

(1. Biological Engineering Institute，Shanxi University，Taiyuan 030006；
2. Biotechnology Research Center，Shanxi Academy of Agricultural Sciences，Taiyuan 030031；
3. Key Laboratory of Crop Gene Resources and Germplasm Enhancement on Loess Plateau， Ministry of Agriculture，Taiyuan 030031)

Received:2013-10-11 Published:2014-04-29 Online:2014-04-29

摘要/Abstract

摘要： 采用花青素调控因子C1/Bperu分别与玉米胚特异性启动子Glb1和组成型启动子CaMV35S构建成植物转化载体pGlb1CB和p35SCB，并利用基因枪转化方法，将重组表达载体转入玉米幼胚。显微观察结果证实，这两个载体均能在玉米幼胚细胞中瞬时表达。用花青素作为标记基因不仅可以在一定程度上减少公众对转基因生物安全性方面的担忧，而且可以帮助直观地从转化当代和后代种子中通过颜色标记筛选到转化籽粒，从而可以大大简化筛选程序，提高效率，节约检测成本。

关键词: 花青素, 直观标记, 载体构建, 瞬时表达, 玉米幼胚

Abstract: Anthocyanin regulatory factor C1/Bperu genes are constructed respectively with a maize embryo specific promoter GLB1 and a constitutive promoter CaMV35S into plant transformation vectors pGlb1CB and p35SCB, and the recombinant expression vectors were introduced into maize embryos by particle bombardment. Microscopic observation confirmed that the two vectors were actively expressed in maize immature embryo cells. The use of anthocyanins as a marker gene not only could reduce the safety concerns from the public over GMO substantially, but could also be used as a visual indicator for the selection of transgenic plants, which could greatly simplify the screening procedures, improve the efficiency and reduce the costs.

Key words: Anthocyanins, Visual marker, Vector construction, Transient expression, Maize immature embryo

赵欣梅, 吴树彪, 王亦学, 崔贵梅, 王晓清, 杜建中, 王小丽, 尚勇进, 孙毅. 花青素合成调控转录基因作为直观标记的载体构建及其在玉米幼胚中的瞬时表达[J]. 生物技术通报, 2014, 0(4): 77-82.

Zhao Xinmei, Wu Shubiao, Wang Yixue, Cui Guimei, Wang Xiaoqing, Du Jianzhong, Wang Xiaoli, Shang Yongjin, Sun Yi,. The Vector Construction of Anthocyanins as a Visual Marker Gene and Its Transient Expression in Maize Immature Embryos[J]. Biotechnology Bulletin, 2014, 0(4): 77-82.

参考文献

[1] 程志强, 陈旭君, 郭泽建, 李德葆.转基因植物中抗生素抗性基因的安全性评价[J].生命科学, 2002, 14(1)：14-16.
[2] Miki B, McHugh S. Selectable marker genes in transgenic plants：applications, alternatives and biosafety[J]. Journal of Biotechnology, 2004, 107(3)：193-232.
[3] 亓燕红.彩色玉米籽粒中参与花青素合成基因的表达分析［D].泰安：山东农业大学, 2011.
[4] Doshi KM, Eudes F, Laroche A, Gaudet D. Transient embryo-specific expression of anthocyanin in wheat[J]. Plant, 2006, 42：432-438.
[5] Doshi KM, Eudes F, Laroche A, Gaudet D. Anthocyanin expression in marker free transgenic wheat and triticale embryos[J]. Plant, 2007, 43：429-435.
[6] Zale JM, Agarwal S, Loar S, et al. Evidence for stable transformation of wheat by floral dip in Agrobacterium tumefaciens[J]. Plant Cell Rep, 2009, 28(6)：903-913.
[7] Liu YS, Petolino JF. Pigmented maize seed via tissue-specific expre-ssion of anthocyanin pathway gene transcription factors[J]. Mol Breeding, 2006, 18：57-67.
[8] 宫硖, 杨凤萍, 薛静, 等.花青素合成转录因子基因在玉米中的表达研究：一种新型基因可视化跟踪表达系统[J].科学通报, 2012, 57(24)：2285-2291.
[9] Kortstee AJ, Khan SA, Helderman C, et al. Anthocyanin production as a potential visual selection marker during plant transformation[J]. Transgenic Res, 2011, 20：1253-1264.
[10] 于丽萍, 亓宝秀, 李新征.玉米种子胚特异性启动子emb5的功能分析［D].泰安：山东农业大学, 2010.
[11] Shepherd CT, Scott MP. Construction and evaluation of a maize(Zea mays)chimaeric promoter with activity in kernel endosperm and embryo[J]. Biotechnology, 2009, 52：233-243.
[12] Liu SQ, Kriz A. Tissue-specific and ABA-regulated Maize Glbl gene expression in transgenic tobacco[J]. Plant Cell Reports, 1996, 16：158-162.
[13] Liu YS, Petolino JF. Pigmented maize seed via tissue-specific expression of anthocyanin pathway gene transcription factors[J]. Mol Breeding, 2006, 18：57-67.
[14] 赵艳, 史岩玲, 钱丹丹, 等.大豆脂肪氧化酶-3(Lox3)启动子的克隆及其瞬时表达分析[J].生物技术通报, 2010(9)：65-69.
[15] Zakharov A, Giersberg M, Hosein F, et al. Seed specific promoters direct gene expression in non-seed tissue[J]. Journal of Experimental Botany, 2004, 55(402)：1463-1471.
[16] Sunilkumar G, Connell JP, Smith CW, et al. Cotton α-globulin promoter：isolation and functional characterization in transgenic cotton, Arabidopsis, and tobacco[J]. Transgenic Research, 2002, 11：347-359.
[