Abiotic stresses, such as salinity, greatly threaten the growth and productivity of plants. Rice (Oryza sativa L.) is one of the most important food crops, as well as a monocot model for genomic research. To obtain a global view of the molecular response to salinity stress, we conducted a leaf transcriptome analysis on rice seedlings. Two cultivars of rice subspecies indica, including the salt-tolerant genotype Xian156 and the salt-sensitive genotype IR28, were used in the present study. Eighteen RNA libraries were obtained from these two genotypes at three timepoints (0 h, 48 h and 72 h) after applying salinity stress. We obtained the reference-guided assembly of the rice transcriptome, which resulted in 1,375 novel genes, including 1,371 annotated genes. A comparative analysis between genotypes and time points showed 5,273 differentially expressed genes (DEGs), of which 286 DEGs were only found in the tolerant genotype. The Disease resistance response protein 206 and TIFY 10 A were differentially expressed, which were validated by quantitative real-time PCR. The differentially expressed genes identified through the mRNA transcriptome, along with the structure, provide a revealing insight into rice molecular response to salinity stress and underlie the salinity tolerance mechanism between genotypes.
Background Isatis indigotica, the source of the traditional Chinese medicine Radix isatidis (Ban-Lan-Gen), is an extremely important economical crop in China. To facilitate biological, biochemical and molecular research on the medicinal chemicals in I. indigotica, here we report the first I. indigotica transcriptome generated by RNA sequencing (RNA-seq).ResultsRNA-seq library was created using RNA extracted from a mixed sample including leaf and root. A total of 33,238 unigenes were assembled from more than 28 million of high quality short reads. The quality of the assembly was experimentally examined by cDNA sequencing of seven randomly selected unigenes. Based on blast search 28,184 unigenes had a hit in at least one of the protein and nucleotide databases used in this study, and 8 unigenes were found to be associated with biosynthesis of indole and its derivatives. According to Gene Ontology classification, 22,365 unigenes were categorized into 48 functional groups. Furthermore, Clusters of Orthologous Group and Swiss-Port annotation were assigned for 7,707 and 18,679 unigenes, respectively. Analysis of repeat motifs identified 6,400 simple sequence repeat markers in 4,509 unigenes.ConclusionOur data provide a comprehensive sequence resource for molecular study of I. indigotica. Our results will facilitate studies on the functions of genes involved in the indole alkaloid biosynthesis pathway and on metabolism of nitrogen and indole alkaloids in I. indigotica and its related species.
BackgroundIsatis indigotica, a traditional Chinese medicine, produces a variety of active ingredients. However, little is known about the key genes and corresponding expression profiling involved in the biosynthesis pathways of these ingredients. Quantitative real-time polymerase chain reaction (qRT-PCR) is a powerful, commonly-used method for gene expression analysis, but the accuracy of the quantitative data produced depends on the appropriate selection of reference genes.ResultsIn this study, the systematic analysis of the reference genes was performed for quantitative real-Time PCR normalization in I. indigotica. We selected nine candidate reference genes, including six traditional housekeeping genes (ACT, α-TUB, β-TUB, UBC, CYP, and EF1-α), and three newly stable internal control genes (MUB, TIP41, and RPL) from a transcriptome dataset of I. indigotica, and evaluated their expression stabilities in different tissues (root, stem, leaf, and petiole) and leaves exposed to three abiotic treatments (low-nitrogen, ABA, and MeJA) using geNorm, NormFinder, BestKeeper, and comprehensive RefFind algorithms. The results demonstrated that MUB and EF1-α were the two most stable reference genes for all samples. TIP41 as the optimal reference gene for low-nitrogen stress and MeJA treatment, while ACT had the highest ranking for ABA treatment and CYP was the most suitable for different tissues.ConclusionsThe results revealed that the selection and validation of appropriate reference genes for normalizing data is mandatory to acquire accurate quantification results. The necessity of specific internal control for specific conditions was also emphasized. Furthermore, this work will provide valuable information to enhance further research in gene function and molecular biology on I. indigotica and other related species.Electronic supplementary materialThe online version of this article (10.1186/s12867-019-0126-y) contains supplementary material, which is available to authorized users.
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