以甘露糖作为筛选剂的棉花遗传转化

棉花学报 Cotton Science 2005,17(1):3-7

以甘露糖作为筛选剂的棉花遗传转化
岳建雄¹，孟钊红²，张炼辉³，韩少杰¹，林亲铁¹，王巍¹
(1.广东省工程技术研究所，广州 510400；2．山西省农科院棉花研究所，运城 044000；
3．新加坡分子农业生物学院, 新加坡117604）

摘要：以 pmi基因作为筛选标记基因，以甘露糖作为筛选剂，通过农杆菌介导法将GFP基因导入棉花细胞并得到再生植株，经过PCR检测、Southern杂交证实外源基因已经整合到棉花基因组中，West bloting与荧光显微镜检测证明GFP基因得到表达。本文研究讨论了甘露糖作为棉花转化细胞的筛选剂在农杆菌介导的转化中的应用浓度及方法，即：甘露糖的筛选浓度在30～50 g·L^-1之间，在愈伤组织诱导初期适当低一点，随着愈伤组织的生长而加大筛选浓度。由于甘露糖不利于再生胚的分化，当愈伤组织转入分化培养基时，要以葡萄糖代替甘露糖。
关键词：棉花；转基因；甘露糖
中图分类号：S562.035.3 文献标识码：A
文章编号：1002-7807（2005）01-0003-05 ［全文，2416KB］

参考文献：
［1］JOERSBO M. Advances in the selection of transgenic plants using non-antibiotic marker genes［J］. siogia plantarum , 2001, 111: 269-272.
［2］JANET Reed, Laura Privalle, Powell M L, et al. Phosphomannose isomerase: an efficient selectable marker for plant transformation［J］. In vitro cell biol-Plant,2001, 37: 127-132.
［3］REED J N,Chang Y-F,McNamara D D, et al. High frequency transformation of wheat with the selectable marker mannose-6-phosphate isomerase(PMI)［M］. In vitro,1999, 35:57-A abstract P-1079.
［4］WANG A S,Evans R A, Altendorf P R, et al. A mannose selection system for production of fertile transgenic maize plants from protoplasts［J］. Plant Cell Reports 2000,19:654-660.
［5］NEROTTO D, Jolley M, Beer S, et al . The use of phosphomannose-isomerase as a selectable marker to recover transgenic maize plants (Zea may L.) via Agrobacterium transformation［J］. Plant cell reports 2000,19: 798-803.
［6］ZHANG P , Puonti-Kaerlas J. PIG-mediated cassava transformation using positive and negative selection［J］. Plant cell reports , 2000, 19: 1041-1048.
［7］ZHANG P, Potrykus I, Johanna P K. Efficient production of transgenic cassava using negative and positive selection［J］. Transgenic research, 2000, 9: 405-415.
［8］PAOLA Lucca , Xu dong Ye . Ingo Potrykus, Effective selection and regeneration of transgenic rice plants with mannose as selective agent［J］. Molecular Breeding, 2001, 7: 43-49.
［9］WRIGHT M, Dawson J , Dunder E et al , Efficient biolistic transformation of maize (Zea mays L.) and wheat (Triticum aestivum L.) using the phosphomannose isomerase gene, pmi, as the selectable marker［J］. Plant cell reports, 2001,20: 429-436.
［10］JOERSBO M, Jorn D M, Janne B. Relationship between promoter strength and transformation frequencies using mannose selection for the production of transgenic sugar bee［J］. Molecular Breeding, 2000, 6: 207-213.
［11］JOERSBO M, Iain D, Jette K, et al. Analysis of mannose selection used fortransformation of sugar beet［J］. Molecular Breeding,1998, 4: 111-117.
［12］WANG Z B, Guo S D. Expressin of two insect-resistant genes cryIA(b&c)GNA in transgenic tobacco plants results in added protection sgainst both cotton bollworm and aphids［J］. Chinese science bulletin, 1999,44(22): 2051-2058.
［13］JOERSBO M, Petersen S G, Okkels F T, Parameters interacting with mannose selection employed for the production of transgenic sugar beet［J］. Physiologia plantarum, 1999,105(1): 109-115.
［14］SHEU-HWA C-S,Lewis D H, Walker DA. Stimulation of photosynthetic starch formation by sequestration of cytoplasmic orthophosphate［J］. New phytologist,1975,74:383-392.
［15］岳建雄，张慧军，张炼辉. 以对潮霉素抗性为筛选标记的棉花遗传转化［J］.棉花学报, 2002,14（4）：195-199.
［16］SOUTHERN E M. Detection of specific sequences among DNA fragments separated by gel electrophoresis ［J］.J Mol Biol,1975,98:503.