An efficient protocol for the establishment of transgenic high hariy root tea plant (Camellia sinensis) system induction infected by Agrobacterium rhizogenes is reported using two wild type strains of A. rhizogenes, three different co-cultivation media and six different explants. The highest induction frequency was obtained from stem segments of aseptic seedlings infected by A. rhizogenes and co-cultivated on YMB solid media with 100βmmol/L AS for 2βd. The hairy roots grew rapidly on LG0 medium without phytohormone and produced a plenty of branches and root hairs. PCR confirmed that rolA, rolB and rolC have inserted into the genomic DNA of hairy root induced by wild type A. rhizogenes. Hairy roots were also induced from explants of tea plant infected by A. rhizogenes 15834 carrying pCAMBIA2301 binary vector harboring Bt gene, transgenic root were confirmed by PCR and GUS histochemical staining that the Bt exogenous gene was inserted and expressed.
ZHANG Guang-hui
,
LIANG Yue-rong
,
LU Jian-liang
. Agrobacterium Rhizogenes-Mediated High Frequency Hairy Root Induction and Genetic Transformation in Tea Plant[J]. Journal of Tea Science, 2006
, 26(1)
: 1
-10
.
DOI: 10.13305/j.cnki.jts.2006.01.001
[1] 王丽鸳, 成浩, 周健. 茶树DNA分子标记及基因工程研究进展[J]. 茶叶科学, 2004, 24(1): 12~17.
[2] Mondal T K, Bhattacharya A, Laxmikumaran M, et al. Recent advances of tea (Camellia sinensis) biotechnology[J]. Plant Cell, Tissue and Organ Culture, 2004, 76: 195~254.
[3] 吴姗, 粱月荣, 陆建良, 等. 茶树农杆菌转化系统和基因枪转化系统的优化[J]. 茶叶科学, 2003, 23(1): 6~10.
[4] 王关林, 方宏筠. 植物基因工程(第二版). 北京:科学出版社, 2002.
[5] Kato M.Regeneration of plantlets from tea stem callus[J]. Japan J Breed, 1985, 35: 317~322.
[6] 中村順行. チャの茎切片培養における不定芽分化とその品種間差異[J]. 茶業研究報告, 1989, 70: 41~49.
[7] Das S, Jha T B, Jha S.Organogenesis and regeneration from pigmented callus in Camellia sinensis(L.) O. Kuntze cv. Nandadevi, an elite Darjeeling tea clone[J]. Plant Science, 1996, 121: 207~211.
[8] 青島洋一. チャ培養植物体茎切片からの不定芽分化率の向上[J]. 茶業研究報告,1998, 87(別冊): 44~45.
[9] Mondal T K, Bhattacharya A, Ahuja P S.Transgenic tea Camellia sinensis (L.) O. Kuntze cv. Kangra Jat plants obtained by Agrobacterium-mediated transformation of somatic embryos[J]. Plant Cell Reports, 2001,20:712~720.
[10] 张广辉, 梁月荣, 吴颖. 咖啡因生物合成研究进展及在茶树育种中的应用[J]. 茶叶,2005, 31(1): 18~23.
[11] Kato M.Somatic embryogenesis from immature leaves of in vitro grown tea shoots[J]. Plant Cell Reports, 1996,15:920~923.
[12] 王毅军, 严学成, 暨淑仪, 等. 茶(Camellia sinensis Kuntze)离体培养体细胞胚胎发生的组织细胞学研究[J]. 热带亚热带植物学报, 1994, 2(2): 47~51.
[13] Zehra M, Banerjee S, Mathur A K, et al. Induction of hairy roots in tea (Camellia sinensis L.) using Agrobacterium rihizogenes[J]. Curr Sci, 1996, 70: 84~86.
[14] 奚彪, 刘祖生, 梁月荣, 等. 发根农杆菌介导的茶树遗传转化[J]. 茶叶科学, 1997, 17(增刊): 155~156.
[15] 高秀清, 成浩, 牛爱军. 茶树毛状根的诱导[J]. 特产研究, 2004, 26(2): 30~32.
[16] 周达锋, 卜学贤, 陈维伦. 发根农杆菌Ri质粒的分子生物学及其应用前景[J]. 植物学通报, 1993, 10(2): 24~34.
[17] 梁机, 陈晓阳, 林善枝, 等. 发根农杆菌Ri质粒rol基因研究进展及在林木改良上的应用[J]. 植物学通报, 2002, 19(6): 650~658.
[18] Torrgrosa L, Bouquet A.Agrobacterium rhizogenes and A. tumefaciens co-transformation to obtain grapevine hairy roots producing the coat protein of grapevine chrome mosaic nepovirus. Plant Cell, Tissue Organ culture, 1997, 49:53~62.
[19] Edwards K,Johnstone C,Thompson C.A simple and rapid method for preparation of plant genomic DNA for PCR analysis.Nucleic Acids Research, 1991, 19(6): 1349.
[20] 中村順行. 2)チャの組織培養を利用した種苗大量増殖法.茶業研究報告, 1991, 74:31~38.
[21] Detlef W, Jane G.拟南芥实验手册(英文影印版). 北京:化学工业出版社, 2004.
[22] 骆颖颖, 粱月荣. Bt基因表达载体的构建及对茶树遗传转化的研究[J]. 茶叶科学, 2000, 20(2): 141~147.
[23] Rurb S, Hensgens L A M. Improved histochemical staining for β-D-Glucuronidase activity in monocotyledonous plants[J]. Rice Genetics Newsletter, 1989, 6: 168~169.
[24] Biao X, Kuboi T, Xu J, Rongyan B. Effect of polyphenol compounds on tea plant transformation. Am. Soc. Plant Physiol, 1998. Abstr. pp314
[25] Subramanian S, Graham M Y, Yu O, et al. RNA interference of soybean isoflavone synthase genes leads to silencing in tissues distal to the transformation site and to enhanced susceptibility to phytophthora sojae[J]. Plant Physiol. 2005, 137(4): 1345~1453.
浙公网安备 33019902000101号 
