Guo, Wanli, Li, Yidan, Gong, Lei, Li, Fengxia, Dong, Yingshan, and Liu, Bao
Robinia ambiguavar. idahoensis, presumably originated from interspecific hybridization of R. pseudoacaciaL. and R. hispidaL., is a multipurpose tree. Several reports have showed that in vitromicropropagation is a feasible method to produce large quantities of ‘clonal’ plants from R. pseudoacacia, however, no information is available on micropropagation of R. ambiguaor the other assumed parental species, R. hispida. Here, we report on a tissue culture system for efficient micropropagation of R. ambiguaplants by enhanced branching of axillary buds taken from a single branch of a donor tree. The culture system consists of sequential use of three media, namely, the bud-induction medium (MS medium supplemented with 0.8–1.4 mg l−16-BA, 0.05–0.08 mg l−1NAA and 0.07–0.1 mg l−1GA), elongation medium (MS medium added with 0.35–0.5 mg l−16-BA, 0.05–0.08 mg l−1NAA and 0.07–0.1 mg l−1GA) and root-induction medium (1/4 MS medium fortified with 1.7–2.5 mg l−1IAA and 0.1–0.5 mg l−1IBA). In addition, we investigated the genetic stability (relative to the donor plant) of a sample of 41 morphologically normal plants randomly taken from ca. 13,000 micropropagated plants, by using the inter-simple sequence repeat (ISSR) marker with 32 selected primers. We found that of the 226 reproducible bands scored, 24 were polymorphic (10.62%), thus pointing to the occurrence, though at a relatively low level compared with an earlier study on R. pseudoacacia, of genomic variation in these micropropagated plants. Further sequencing on seven loci underlying the variations showed that two had significant homology to known or predicted plant genes.Robinia ambiguavar. idahoensis, presumably originated from interspecific hybridization of R. pseudoacaciaL. and R. hispidaL., is a multipurpose tree. Several reports have showed that in vitromicropropagation is a feasible method to produce large quantities of ‘clonal’ plants from R. pseudoacacia, however, no information is available on micropropagation of R. ambiguaor the other assumed parental species, R. hispida. Here, we report on a tissue culture system for efficient micropropagation of R. ambiguaplants by enhanced branching of axillary buds taken from a single branch of a donor tree. The culture system consists of sequential use of three media, namely, the bud-induction medium (MS medium supplemented with 0.8–1.4 mg l−16-BA, 0.05–0.08 mg l−1NAA and 0.07–0.1 mg l−1GA), elongation medium (MS medium added with 0.35–0.5 mg l−16-BA, 0.05–0.08 mg l−1NAA and 0.07–0.1 mg l−1GA) and root-induction medium (1/4 MS medium fortified with 1.7–2.5 mg l−1IAA and 0.1–0.5 mg l−1IBA). In addition, we investigated the genetic stability (relative to the donor plant) of a sample of 41 morphologically normal plants randomly taken from ca. 13,000 micropropagated plants, by using the inter-simple sequence repeat (ISSR) marker with 32 selected primers. We found that of the 226 reproducible bands scored, 24 were polymorphic (10.62%), thus pointing to the occurrence, though at a relatively low level compared with an earlier study on R. pseudoacacia, of genomic variation in these micropropagated plants. Further sequencing on seven loci underlying the variations showed that two had significant homology to known or predicted plant genes.