Variation at the transcriptional level among Chinese natural populations of Arabidopsis thaliana in response to cold stress

The Arabidopsis 25K GeneChip (ATH1, Affymetrix) was used to make a survey of the variation of the transcriptional profiles among 5 Chinese natural populations of Arabidopsis thaliana under cold treatment. In normal growth condition, the expression level of 2.26% (513 genes in the population from Jiujiang, Jiangxi, JXjjx) to 6.52% (1482 genes in the population from Tongliang, Chongqing, CQtlx) genes was 2-fold higher than that of Col ecotype. Under cold treatment, the expression of 12.84% (2920 genes in the population from Chenggu, Shaanxi, SXcgx) to 19.46% (4426 genes in the population from Qinghe, Xinjiang, XJqhx) genes was up- or down-regulated by at least two-fold that of their controls. In general, most of up-regulated genes might be the genes essential for plant surviving at low temperature, such as genes in CBF pathway and the genes responsible for synthesizing molecules accumulated for cold tolerance. However, each natural population had some specific genes induced under cold treatment. The data indicated that some of the cold-responding genes were differentiated among the populations distributed in the natural habitats with different climate conditions. CBF3, one of the key tran-scription factor genes in cold responding pathway, showed significant differences in expression among populations. The sequence analysis indicated that the changes in its regulation region caused the dramatic difference in the expression pattern. Further studies on the correlation of the function of the differentially expressed genes and the cold tolerance in different populations may provide some new insight into the molecular mechanism of adaptation to local environment in Arabidopsis thaliana in China.

Construction of chimeric inducible promoters by elicitors of rice fungal blast pathogen and their expression in transgenic rice

The promoter fragments of wheat GstA1 and potato Gst1 have been amplified by PCR, cloned and fused respectively to the minimal promoter sequence of rice actin gene (Act1)) and its 5’ untranslated leader sequence together with GUS. The constructs with 2 chimeric promoters (WGA and PGA) have been transferred into rice in order to analyze their inducibility patterns in transgenic rice plants. The results show that: WGA and PGA are both inducible by elicitors of Pyricularia oryzae in transgenic rice cells; the intron I of rice Act1 gene is important for the heterogenic expression of monocot and dicot promoter elements in rice; and the Act1 minimal promoter and its 5’untranslated leader sequence produced low level background expression in rice.

Molecular evolution of the exon 2 of CHS genes and the possibility of its application to plant phylogenetic analysis

The exon 2 of chalcone synthase (CHS) gene is relatively conserved during evolution. In this study, three exon 2 fragments from two species in gymnosperm (Cycas panzhihuaensis, Ginkgo biloba ) and seven from four species in angiosperm (Magnolia denudata, Salix babylonica, Nymphaea tetragona, Camellia japonica) have been amplified by PCR from genomic DNA and sequenced. Together with other 73 sequences of CHS collected from EMBL database and literature, these sequences, which embrace 19 families of gymnosperm and angiosperm, have been analyzed for their phylogenetic relations by parsimony method. The result indicated that sequences from the same systematic family usually grouped together except those from Theaceae, Magnoliaceae and Nymphaeaceae. The relative rate test revealed the rate heterogeneity of CHS genes among the families. For the nucleotide substitution the sequences from Asteraceae and Solanaceae evolve faster than those from the other families analyzed while the sequences from Poaceae,

Expression and inhibitory activity analysis of a 25-kD Bowman-Birk protease inhibitor in rice

Rice Bowman-Birk inhibitors (RBBI), with one (8 kD) or two homologous domains (16 kD), were found to be effective trypsin inhibitors in vitro. In this study, we demonstrate that the 25-kD protein corresponding to the three-domain RBBI indeed ex- ists in rice in planta, and that the RBBIs are regulated by development and wounding. We also found by inhibitory activity assay that the 3:13 disulfide bond, but not the 4:5 disulfide bond, suppresses the tryp- sin-inhibitory activity, and the D3 domain of RBBI3-1 has no inhibitory activity against trypsin, chymotryp- sin, paparin or subtilisin. Mutation analyses showed that conversion from Lys to Leu or Tyr in the N-terminal P1 site in D1 domain did not create chy- motrypsin-inhibitory activity, suggesting that the structure of the reactive loop in D1 domain hinder the new inhibitory specificity at P1 site, and the chy- motrypsin-inhibitory activity might need the participa- tion of other structures, e.g. 3:13 disulfide bond.

Cloning and expression of a cDNA encoding ribosomal protein S4 from Rice (Oryza sativa)

A cDNA clone, pS4, has been isolated from a cDNA library prepared from rice anthers of about 1.0 mm in length. DNA sequence analysis and database search show that the cDNA encodes a protein which is highly homologous to eukaryotic 80S ribosomal protein subunit 4 (S4). Northern hybridization indicates that this gene expresses in all tissues analyzed although the expression level varies and it cannot be induced by mechanical wounding in leaves. Southern blot analysis demonstrates that this rice S4 gene is from a multigene