[Cloning, subcellular lacalization and expression analysis of chloroplast-targeted Obg gene in Cyathula officinalis].

According to ObgC gene sequences from Cyathula officinalis genomic data, the specific primers were designed, and a full-length CoObgC cDNA (2 226 bp) was obtained by polymerase chain reaction (PCR) and rapid amplification of cDNA ends (RACE) methord. Sequence alignment showed that CoObgC gene contained a 1 818 bp open reading frame (ORF) encoding 605 amino acids. Sequence analysis predicted that molecular weight of CoObgC protein was about 66.39 kDa, the academic isoelectric point was 5.35, and the protein was stable protein. Then multiple sequence alignment was applied to construct phylogenetic tree. The real-time fluorescence quantification PCR (RT-qPCR) demonstrated that a high expression level in leaf, followed by root and flower, the low transcription was in stem. The recombinant vector pCABIA2300-CoObgC was constructed and introduced into tobacco epidermal cells by agrobacterium-mediated transformation, green fluorescence was tested and targeted to chloroplast under a laser scanning confocal microscope. These findings will be helpful to lay a foundation for studying the structure and function of CoObgC gene, and elucidating C. officinalis molecular biology experiment.

[Physiological and biochemical change of Paris seed in after-ripening during variable temperature stratification].

In order to explore the dormancy physiological and biochemical mechanism of Paris seeds, the seed embryo growth courses, and the dynamic change of 5 enzymes, include SOD, POD, CAT, MDH, G-6-PDH were measured during variable temperature stratification. The results indicated that Paris seeds embryo grew quickly after 40 d in warm-stratification (18 ± 1) °C, at the meantime the metabolic activity was significantly strengthened. These facts showed that Paris seeds turned into physiological after-ripening process. After 60-80 d, the morphological embryo after-ripping process basically completed, and the following cold-stratification (4 ± 1) °C furthered Paris seed to finish physiological after-ripening. After 40 d, the activity of MDH decreased while G-6-PDH increased significantly. This showed that the main respiratory pathway of seed changed from TCA to PPP, which benifited breaking seed dormancy. In the whole period of stratification process, the activity variation of SOD and CAT was insignificantly and the activity of POD was enhanced significantly after shifting the seed in cold stratification process. This showed that SOD, CAT had no direct effects on breaking Paris seed dormancy but keeping the seed vigor, while the POD might involve in the process of Paris seed dormancy breaking.