Genome-Wide Transcriptomic Identification and Functional Insight of Lily WRKY Genes Responding to Botrytis Fungal Disease.

Genome-wide transcriptome analysis using RNA-Seq of Lilium longiflorum revealed valuable genes responding to biotic stresses. WRKY transcription factors are regulatory proteins playing essential roles in defense processes under environmental stresses, causing considerable losses in flower quality and production. Thirty-eight WRKY genes were identified from the transcriptomic profile from lily genotypes, exhibiting leaf blight caused by Botrytis elliptica. Lily WRKYs have a highly conserved motif, WRKYGQK, with a common variant, WRKYGKK. Phylogeny of LlWRKYs with homologous genes from other representative plant species classified them into three groups- I, II, and III consisting of seven, 22, and nine genes, respectively. Base on functional annotation, 22 LlWRKY genes were associated with biotic stress, nine with abiotic stress, and seven with others. Sixteen unique LlWRKY were studied to investigate responses to stress conditions using gene expression under biotic and abiotic stress treatments. Five genes-LlWRKY3, LlWRKY4, LlWRKY5, LlWRKY10, and LlWRKY12-were substantially upregulated, proving to be biotic stress-responsive genes in vivo and in vitro conditions. Moreover, the expression patterns of LlWRKY genes varied in response to drought, heat, cold, and different developmental stages or tissues. Overall, our study provides structural and molecular insights into LlWRKY genes for use in the genetic engineering in Lilium against Botrytis disease.

Transcriptome Analysis of Diurnal Gene Expression in Chinese Cabbage.

Plants have developed timing mechanisms that enable them to maintain synchrony with daily environmental events. These timing mechanisms, i.e., circadian clocks, include transcriptional/translational feedback loops that drive 24 h transcriptional rhythms, which underlie oscillations in protein abundance, thus mediating circadian rhythms of behavior, physiology, and metabolism. Circadian clock genes have been investigated in the diploid model plant Arabidopsis thaliana. Crop plants with polyploid genomes-such as Brassica species-have multiple copies of some clock-related genes. Over the last decade, numerous studies have been aimed at identifying and understanding the function of paralogous genes with conserved sequences, or those that diverged during evolution. Brassicarapa's triplicate genomes retain sequence-level collinearity with Arabidopsis. In this study, we used RNA sequencing (RNAseq) to profile the diurnal transcriptome of Brassicarapa seedlings. We identified candidate paralogs of circadian clock-related genes and assessed their expression levels. These genes and their related traits that modulate the diurnal rhythm of gene expression contribute to the adaptation of crop cultivars. Our findings will contribute to the mechanistic study of circadian clock regulation inherent in polyploidy genome crops, which differ from those of model plants, and thus will be useful for future breeding studies using clock genes.

Generation and analysis of expressed sequence tags (ESTs) of Camelina sativa to mine drought stress-responsive genes.

Camelina sativa is an oil-producing crop belonging to the family of Brassicaceae. Due to exceptionally high content of omega fatty acid, it is commercially grown around the world as edible oil, biofuel, and animal feed. A commonly referred 'false flax' or gold-of-pleasure Camelina sativa has been interested as one of biofuel feedstocks. The species can grow on marginal land due to its superior drought tolerance with low requirement of agricultural inputs. This crop has been unexploited due to very limited transcriptomic and genomic data. Use of gene-specific molecular markers is an important strategy for new cultivar development in breeding program. In this study, Illumina paired-end sequencing technology and bioinformatics tools were used to obtain expression profiling of genes responding to drought stress in Camelina sativa BN14. A total of more than 60,000 loci were assembled, corresponding to approximately 275 K transcripts. When the species was exposed to 10 kPa drought stress, 100 kPa drought stress, and rehydrated conditions, a total of 107, 2,989, and 982 genes, respectively, were up-regulated, while 146, 3,659, and 1189 genes, respectively, were down-regulated compared to control condition. Some unknown genes were found to be highly expressed under drought conditions, together with some already reported gene families such as senescence-associated genes, CAP160, and LEA under 100 kPa soil water condition, cysteine protease, 2OG, Fe(II)-dependent oxygenase, and RAD-like 1 under rehydrated condition. These genes will be further validated and mapped to determine their function and loci. This EST library will be favorably applied to develop gene-specific molecular markers and discover genes responsible for drought tolerance in Camelina species.

Expression and characterization of codon-optimized carbonic anhydrase from Dunaliella species for CO(2) sequestration application.

Carbonic anhydrases (CAs) have been given much attention as biocatalysts for CO(2) sequestration process because of their ability to convert CO(2) to bicarbonate. Here, we expressed codon-optimized sequence of α-type CA cloned from Dunaliella species (Dsp-aCAopt) and characterized its catalyzing properties to apply for CO(2) to calcite formation. The expressed amount of Dsp-aCAopt in Escherichia coli is about 50 mg/L via induction of 1.0 mM isopropyl-ß-D-thiogalactopyranoside at 20 °C (for the case of intact Dsp-aCA, negligible). Dsp-aCAopt enzyme shows 47 °C of half-denaturation temperature and show wide pH stability (optimum pH 7.6/10.0). Apparent values of K (m) and V (max) for p-nitrophenylacetate substrate are 0.91 mM and 3.303 × 10(-5) µM min(-1). The effects of metal ions and anions were investigated to find out which factors enhance or inhibit Dsp-aCAopt activity. Finally, we demonstrated that Dsp-aCAopt enzyme can catalyze well the conversion of CO(2) to CaCO(3), as the calcite form, in the Ca(2+) solution [8.9 mg/100 µg (172 U/mg enzyme) with 10 mM of Ca(2+)]. The obtained expression and characterization results of Dsp-aCAopt would be usefully employed for the development of efficient CA-based system for CO(2)-converting/capturing processes.