Selection of non-branching lines induced by introducing Ls-like cDNA into Chrysanthemum ( Dendranthema × grandiflorum (Ramat.) Kitamura) “Shuho-no-chikara”

Abstract

A partial Ls-like cDNA (525bp) was isolated from Chrysanthemum “95B1-45” line, which exhibited a non-branching characteristic. The fragments showed homology with lateral suppressor ( Ls) partial cDNA from tomato that stimulated the initiation of axillary meristems. After the fragments was constructed with sense and antisense versions, these constructs were employed in Agrobacterium-mediated transformation of Chrysanthemum ( Dendranthema × grandiflorum (Ramat.) Kitamura) “Shuho-no-chikara”. The leaf segments (5 mm × 5 mm) from in vitro shoots were inoculated with Agrobacterium cultures for 10 min and co-cultured with Agrobacterium tumefaciens for three days at 25 °C. Transgenic plants were obtained only in Agrobacterium C58C1 containing pCAMBIA2301 constructed with Ls-like partial cDNA in antisense version. Forty shoots (1 was in positive control and 39 in Ls-antisense version) in Agrobacterium C58C1 were confirmed as putative transformants in PCR analysis with nptII primer. In the Southern analysis, results showed that 35 of the 39 shoots integrated npt II gene were confirmed to contain Ls-like partial cDNA in antisense version. Therefore, the 35 transformants were multiplied and planted in green house for integrated Ls-like partial cDNA expression. One (2.9%) among the 36 transformed lines showed lateral bud inhibition of more than 40%, with another one (2.9%) in lateral bud inhibition of 20–30%, and the remaining ones in less than 20%. However, in the second investigation (in a lateral branch in October 29), nine lines belonged to the range of 20–30% lateral bud inhibition and the remaining ones were in the range of less than 20%. The expression of non-branching characteristics showed significant difference even in the same line according to season.