农杆菌介导的遗传转化已被广泛应用于植物转基因研究。作为外源基因的载体, 农杆菌T-DNA片段在植物基因组中的整合方式, 不仅影响外源基因的整合效率及稳定性, 还会影响外源基因的表达特性。文章就农杆菌介导的T-DNA整合的两种主要模式、规律及相关研究手段进行综述, 为农杆菌介导的转基因及T-DNA插入突变等相关研究提供借鉴。
The genetic transformation mediated by Agrobacterium tumefaciens has been widely applied to research of transgenic plants. As the vector of the exotic genes, the integration patterns of T-DNA fragments affects not only transformation efficiency and stability, but also expression properties of the transgenes. This review summaries the two major patterns and the rules of T-DNA integration in Agrobacterim-mediated transformation, rules of T-DNA me-diated by Agrobacterium tumefaciens, as well as research tools for flanking sequence amplification. It is attempted to provide references for researches on transformation and T-DNA integration mutation mediated by Agrobacterium tumefaciens.
[1] Hamilton CM, Frary A, Lewis C, Tanksley SD. Stable transfer of intact high molecular weight DNA into plant chromosomes. Proc Natl Acad Sci USA, 1996, 93(18): 9975-9979.
[2] Zhu CF, Wu JH, He CZ. Induction of chromosomal inversion by integration of T-DNA in the rice genome. J Genet Genomics, 2010, 37(3): 189-196.
[3] Meinke D, Scholl R. The preservation of plant genetic resources. Experiences with Arabidopsis. Plant Physiol, 2003, 133(3): 1046-1050.
[4] Zhu QH, Ramm K, Eamens AL, Dennis ES, Upadhyaya NM. Transgene structures suggest that multiple mechanisms are involved in T-DNA integration in plants. Plant Sci, 2006, 171(3): 308-322.
[5] 杨继芳, 刘明, 安利佳. T-DNA整合的研究进展. 遗传, 2004, 26(6): 991-996.
[6] Nacry P, Camilleri C, Courtial B, Caboche M, Bouchez D. Major chromosomal rearrangements induced by T-DNA transformation in Arabidopsis. Genetics, 149(2): 641-650.
[7] Cáceres M, Ranz JM, Barbadilla A, Long M, Ruiz Z. Generation of a widespread Drosophila inversion by a transposable element. Science, 285(5426): 415-418.
[8] Tzfira T, Li JX, Lacroix B, Citovsy V. Agrobacterium T-DNA integration: molecules and models. Trends Genet, 2004, 20(8): 375-383.
[9] Kim SR, Jeon JS, An G. Development of an efficient inverse PCR method for isolating gene tags from T-DNA insertional mutants in rice. Plant Reverse Genetics, 2011, 678: 139-146.
[10] Zupan J, Muth TR, Draper O, Zambryski P. The transfer of DNA from Agrobacterium tumefaciens into plants: a feast of fundamental insights. Plant J, 2000, 23(1): 11-28.
[11] Kohli A, Leech M, Vain P, Laurie DA, Christou P. Transgene organization in rice engineered through direct DNA transfer supports a two-phase integration mechanism mediated by the establishment of integration hot spots. Proc Natl Acad Sci USA, 1998, 95(12): 7203-7208.
[12] Ziemienowicz A, Tinland B, Bryant J, Gloeckler V, Hohn B. Plant enzymes but not Agrobacterium VirD2 mediate T-DNA ligation in vitro. Mol Cell Biol, 2000, 20(17): 6317-6322.
[13] Kumar S, Fladung M. Gene stability in transgenic aspen (Populus). II. Molecular characterization of variable expression of transgene in wild and hybrid aspen. Planta, 2001, 213(5): 731-740.
[14] Kumar S, Fladung M. Transgene integration in aspen: structures of integration sites and mechanism of T-DNA integration. Plant J, 2002, 31(4): 543-551.
[15] Tinland B. The integration of T-DNA into plant genomes. Trends Plant Sci, 1996, 1(6): 178-184.
[16] Zupan JR, Citovsky V, Zambryski P. Agrobacterium VirE2 protein mediates nuclear uptake of single-stranded DNA in plant cells. Proc Natl Acad Sci USA, 1996, 93(6): 2392-2397.
[17] Lu J, den Dulk-Ras A, Hooykaas PJJ, Glover JNM. Agrobacterium tumefaciens VirC2 enhances T-DNA transfer and virulence through its C-terminal ribbon-helix-helix DNA-binding fold. Proc Natl Acad Sci USA, 2009, 106(24): 9643-9648.
[18] Zhang YG, Zeng FS, Xin Y. Progress on molecular mechanism of T-DNA transport and integration. Acta Genet Sin, 2005, 32(6): 655-665.
[19] Salomon S, Puchta H. Capture of genomic and T-DNA sequences during double-strand break repair in somatic plant cells. EMBO J, 1998, 17(20): 6086-6095.
[20] Zhang J, Guo D, Chang YX, You CJ, Li XW, Dai XX, Weng QJ, Zhang JW, Chen GX, Li XH, Liu HF, Han B, Zhang QF, Wu CY. Non-random distribution of T-DNA insertions at various levels of the genome hierarchy as re-vealed by analyzing 13 804 T-DNA flanking sequences from an enhancer-trap mutant library. Plant J, 2007, 49(5): 947-959.
[21] Muller AE, Atkinson RG, Sandoval RB, Jorgensen RA. Microhomologies between T-DNA ends and target sites often occur in inverted orientation and may be responsible for the high frequency of T-DNA-associated inversions. Plan
