Regeneration and transformation of Eucalyptus camaldulensis
Regeneration and transformation of Eucalyptus camaldulensis
复制标题
赤桉的再生与转化
DOI：
10.1007/s002990050321
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发表时间：
1997
影响因子：
6.2
通讯作者：
Elizabeth S. Dennis
中科院分区：
文献类型：
--
作者：
K. Mullins;Danny J. Llewellyn;V. Hartney;Steven H. Strauss;Elizabeth S. Dennis
已制定了可靠的赤桉试管苗叶片外植体再生方案。在测试的24个克隆中，13个获得了再生，其中6个获得了超过60%的外植体再生。相同的方法也成功地再生了一些微囊藻、嗜铬偃麦草、大麦和边缘偃麦草的无性系，但再生频率较低。将甘蓝叶片外植体与携带卡那霉素抗性基因和报告基因β-葡萄糖醛酸酶(GUS)的根癌农杆菌菌株共培养，然后用9 mg L-1的卡那霉素进行筛选，可以筛选出能在选择培养基上生根的转化芽。通过对不同植物组织中GUS酶的染色、NptII检测以及利用GUS和可选择标记基因的特异探针对分离的DNA进行Southern blotting，证实了这些植物的转化。用5个骆驼蓬克隆和多个根癌农杆菌菌株进行转化，获得了转化组织。然而，只有1个无性系可靠地再生了整株植株。
Reliable regeneration protocols forEucalyptus camaldulensisusing leaf explants from in vitro-grown plants have been developed. Out of the 24 clones tested 13 were regenerated and of these, 6 showed regeneration from more than 60% of the explants. Identical protocols were also successful in the regeneration of some clones ofE.microtheca,E.ochrophloia,E.grandisandE.marginata, but at lower frequencies. Co-cultivation ofE.camaldulensisleaf explants withAgrobacterium tumefaciensstrains carrying a kanamycin resistance gene and the reporter geneβ-glucuronidase (GUS), followed by selection on kanamycin at 9 mg l–1, allowed the selection of transformed shoots that could be rooted on selective media. Transformation of the plants was verified by staining for the GUS enzyme in various plant tissues, NptII assays and by Southern blotting on isolated DNA using specific probes for both the GUS and selectable marker genes. Transformed tissue was obtained with 5 clones ofE.camaldulensistested and a number ofA.tumefaciensstrains. However, only 1 clone regenerated transformed whole plants reliably.