BcMF30a and BcMF30c, Two Novel Non-Tandem CCCH Zinc-Finger Proteins, Function in Pollen Development and Pollen Germination in Brassica campestris ssp. chinensis.

Chinese cabbage (Brassica campestris) is an economically important leaf vegetable crop worldwide. Mounting studies have shown that cysteine-cysteine-cysteine-histidine (CCCH) zinc-finger protein genes are involved in various plant growth and development processes. However, research on the involvement of these genes in male reproductive development is still in its infancy. Here, we identified 11 male fertility-related CCCH genes in Chinese cabbage. Among them, a pair of paralogs encoding novel non-tandem CCCH zinc-finger proteins, Brassica campestris Male Fertility 30a (BcMF30a) and BcMF30c, were further characterized. They were highly expressed in pollen during microgametogenesis and continued to express in germinated pollen. Further analyses demonstrated that both BcMF30a and BcMF30c may play a dual role as transcription factors and RNA-binding proteins in plant cells. Functional analysis showed that partial bcmf30a bcmf30c pollen grains were aborted due to the degradation of pollen inclusion at the microgametogenesis phase, and the germination rate of viable pollen was also greatly reduced, indicating that BcMF30a and BcMF30c are required for both pollen development and pollen germination. This research provided insights into the function of CCCH proteins in regulating male reproductive development and laid a theoretical basis for hybrid breeding of Chinese cabbage.

The zinc-finger transcription factor BcMF20 and its orthologs in Cruciferae which are required for pollen development.

Brassica campestris Male Fertility 20 (BcMF20) is a typical zinc-finger transcription factor that was previously isolated from flower buds of Chinese cabbage (Brassica campestris ssp. chinensis). By applying expression pattern analysis, it can be known that BcMF20 was specifically and strongly expressed in tapetum and pollen, beginning from the uninucleate stage, and was maintained during the mature-pollen stage. As BcMF20 was highly conserved in Cruciferae, it can be indicated that this zinc-finger transcription factor is important during the growth of Cruciferae. In this study, 12 C2H2-type zinc-finger TFs which shared high homology with BcMF20 were found from NCBI via BLAST. A new molecular phylogenetic tree was constructed by the comparison between BcMF20 and these 12 C2H2-type zinc-finger TFs with NJ method. By analyzing this phylogenetic tree, the evolution of BcMF20 was discussed. Then, antisense RNA technology was applied in the transgenesis of Arabidopsis thaliana to get the deletion mutants of BcMF20, so that its function during the pollen development can be identified. The results showed: BcMF20 are in the same clade with three genes from Arabidopsis. The inhibition of BcMF20 expression led to smaller amounts of and lower rate in germination of pollen and lower rate in fruit setting in certain transgenetic plants. This also led to the complete collapse of pollen grains. By SEM and TEM, pollen morphology and anther development processes were observed. In the middle uninucleate microspore stage, a relatively thin or even no primexine was formed in microspores. This may result in the malformation of the pollen wall and finally cause the deformity of pollens. Above all, it can be indicated that BcMF20 may act as a part of regulation mechanisms of TAZ1 and MS1. Together they play a role in a genetic pathway in the tapetum to act on proliferation of tapetal cells and keep the normal development of pollens.

The endophytic bacterium relieved healthy risk of pakchoi intercropped with hyperaccumulator in the cadmium polluted greenhouse vegetable field.

Planting leafy vegetables, especially pakchoi, in cadmium (Cd) polluted farmland is easy to lead to excessive Cd content in edible parts, which results in high risk of food chain. In this study, a field experiment was carried out to study the effects of intercropping of pakchoi with Cd hyperaccumulator Sedum alfredii Hance, and the roles of endophytic bacterium SaMR12 was also investigated. When intercropping with Sedum, the growth of pakchoi was not affected but their Cd concentration and accumulation were significantly increased, while which were obviously decreased by SaMR12 inoculation. After intercropping, the biomass of Sedum was significantly reduced, but their Cd concentration increased. SaMR12 inoculation significantly increased Cd accumulation of Sedum, and which increased to 3 times in Sedum monoculture. Those results showed that although intercropping with hyperaccumulator could lead to higher risk of pakchoi in Cd polluted field, intercropping with SaMR12 inoculated Sedum can decrease Cd concentration of pakchoi and promote Cd absorption of Sedum, which indicated that this endophyte can be made into a microbial inoculum as a soil additive for the safe production of vegetables and the soil Cd pollution remediation.

Molecular cloning and expression analysis of a Cu/Zn SOD gene (BcCSD1) from Brassica campestris ssp. chinensis.

Superoxide dismutases (SODs) are a family of metalloproteins extensively exists in eukaryote, which plays an essential role in stress-tolerance of higher plants. A full-length cDNA encoding Cu/Zn SOD (BcCSD1) was isolated from young seedlings of non-heading Chinese cabbage (Brassica campestris ssp. chinensis) by rapid amplification of cDNA ends (RACE). Bioinformatics analysis revealed that BcCSD1 belonged to the plant SOD super family and had the closest relationship with SOD from Brassica napus. Tissue expression pattern analysis revealed that the BcCSD1 was constitutively expressed in all the tested tissues, and strongest in leaf, moderate in stem, lowest in root. The expression profiles under different stress treatments such as drought, NaCl, high temperature and ABA were also investigated, and the results revealed that BcCSD1 was a stress-responsive gene, especially to ABA. These results provide useful information for further understanding the role of BcCSD1 resistant to abiotic stress in Brassica campestris in the future.

Occurrence of Clubroot on Pak-Choi Caused by Plasmodiophora brassicae.

Clubroot symptoms occurred severely on roots of Pak-Choi (Brassica campestris ssp. chinensis) grown in greenhouses in Gwangju city, Gyeonggi province, Korea in September, 2008. The incidence of the disease symptoms reached as high as 90% in three greenhouses investigated. The root galls collected from the greenhouses were sectioned using a scalpel and observed by light microscope. Many resting spores were found in the cells of the root gall tissues. Suspension of resting spores was prepared from the root galls and inoculated to roots of healthy Pak-Choi plants. Each of five resting spore suspensions caused clubroot symptoms on the roots, which were similar to those observed during the greenhouse survey. Resting spores of the pathogen were observed in the cells of the affected roots. The clubroot pathogen was identified as Plasmodiophora brassicae based on its morphological and pathological characteristics. This is the first report that Plasmodiophora brassicae causes clubroot of Pak-Choi.

Occurrence of Clubroot on Pak-Choi Caused by Plasmodiophora brassicae

Clubroot symptoms occurred severely on roots of Pak-Choi (Brassica campestris ssp. chinensis) grown in greenhouses in Gwangju city, Gyeonggi province, Korea in September, 2008. The incidence of the disease symptoms reached as high as 90% in three greenhouses investigated. The root galls collected from the greenhouses were sectioned using a scalpel and observed by light microscope. Many resting spores were found in the cells of the root gall tissues. Suspension of resting spores was prepared from the root galls and inoculated to roots of healthy Pak-Choi plants. Each of five resting spore suspensions caused clubroot symptoms on the roots, which were similar to those observed during the greenhouse survey. Resting spores of the pathogen were observed in the cells of the affected roots. The clubroot pathogen was identified as Plasmodiophora brassicae based on its morphological and pathological characteristics. This is the first report that Plasmodiophora brassicae causes clubroot of Pak-Choi.