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Using high competent shoot apical meristems of cockscomb as explants for studying function of ASYMMETRIC LEAVES2-LIKE11 (ASL11) gene of Arabidopsis.

Research paper by Shao-Bo SB Sun, Lai-Sheng LS Meng, Xu-Dong XD Sun, Zhen-Hua ZH Feng

Indexed on: 23 Mar '10Published on: 23 Mar '10Published in: Molecular Biology Reports



Abstract

Though shoot apical meristems (SAMs) commonly exhibit low or no competence for transformation, the potent regeneration of this tissue merits further research. Especially, when shoot regeneration is recalcitrant using other tissues as explants, SAM probably is an excellent selection. In cockscomb plants, using SAMs from seedlings obtained from MS medium with 0.5 mg l(-1) 6-BA as explants, high frequency of transformation (approximate 20%) is obtained; whereas control SAMs performed poorly for transformation (approximate 3%). These SAMs are malformed in morphology compared to control SAMs. Further observation found that, in these SAMs, cell proliferation and/or TE formation are seen; which are not found in control SAMs. GUS assays indicated that GUS-positive blue spots at TE zones are obvious; whereas the case was contrary in control SAMs. All these data suggest that cell proliferation and/or TE formation might cause high effective transformation. This transformation system should facilitate the use of this species for studies on gene manipulation and expression. Therefore, we introduced 35S:ASL11-GFP to cockscomb via Agrobacterium tumefaciens. ASYMMETRIC LEAVES2-LIKE11 (ASL11) gene of Arabidopsis is a member of the ASYMMETRIC LEAVES2 (AS2)/LATERAL ORGAN BOUNDARIES (LOB) domain gene family, and its function is largely unclear. By confocal laser scanning microscopy, we found that in most over 35S:ASL11-GFP cockscomb plants, ASL11-GFP fusion protein was in discrete nuclear location. These results indicate that the T-DNA contains within the construct inserted into the host chromosomes in an integral form, and also suggest that ASL11 might be a nuclear protein and function as a potential transcription factor. Moreover, SAMs of the over 35S:ASL11-GFP plants show needle-like patterns that lack organ primordial; suggesting ASL11 might be involved in sustaining indeterminate cell fate of SAMs.