Grant Jan, Dale Tracy, Cooper Pauline
New Zealand Institute for Crop and Food Research Ltd., Christchurch, New Zealand.
Methods Mol Biol. 2006;344:135-41. doi: 10.1385/1-59745-131-2:135.
This chapter describes the transformation of Pinus radiata using organogenic cotyledon explants rather than the more common somatic embryogenesis methods for conifers. The advantages of our method are the year round availability of seed and that over 80% of genotypes can be easily regenerated from the mature cotyledon explants. The transformation efficiency (i.e., the number of transformed shoots regenerated from excised cotyledons) is 1.7% and, as with other Agrobacterium tumefaciens transformation methods, the majority of transgene integrations are single copy. Critical factors for success are survival of the cotyledons, Agrobacterium strain, and selection pressure after cocultivation.
本章描述了利用器官发生的子叶外植体转化辐射松的方法,而不是针叶树更常用的体细胞胚胎发生方法。我们方法的优点是种子全年可得,并且超过80%的基因型能够从成熟子叶外植体轻松再生。转化效率(即从切除的子叶再生的转化芽数)为1.7%,与其他根癌农杆菌转化方法一样,大多数转基因整合为单拷贝。成功的关键因素是子叶的存活、农杆菌菌株以及共培养后的选择压力。