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Full-length transcriptome sequences and the identification of putative genes for flavonoid biosynthesis in safflower.

ABSTRACT: BACKGROUND:The flower of the safflower (Carthamus tinctorius L.) has been widely used in traditional Chinese medicine for the ability to improve cerebral blood flow. Flavonoids are the primary bioactive components in safflower, and their biosynthesis has attracted widespread interest. Previous studies mostly used second-generation sequencing platforms to survey the putative flavonoid biosynthesis genes. For a better understanding of transcription data and the putative genes involved in flavonoid biosynthesis in safflower, we carry our study. RESULTS:High-quality RNA was extracted from six types of safflower tissue. The RNAs of different tissues were mixed equally and used for multiple size-fractionated libraries (1-2, 2-3 and 3-6 k) library construction. Five cells were carried (2 cells for 1-2 and for 2-3 k libraries and 1 cell for 3-6 k libraries). 10.43Gb clean data and 38,302 de-redundant sequences were captured. 44 unique isoforms were annotated as encoding enzymes involved in flavonoid biosynthesis. The full length flavonoid genes were characterized and their evolutional relationship and expressional pattern were analyzed. They can be divided into eight families, with a large differences in the tissue expression. The temporal expressions under MeJA treatment were also measured, 9 genes are significantly up-regulated and 2 genes are significantly down-regulated. The genes involved in flavonoid synthesis in safflower were predicted in our study. Besides, the SSR and lncRNA are also analyzed in our study. CONCLUSIONS:Full-length transcriptome sequences were used in our study. The genes involved in flavonoid synthesis in safflower were predicted in our study. Combined the determination of flavonoids, CtC4H2, CtCHS3, CtCHI3, CtF3H3, CtF3H1 are mainly participated in MeJA promoting the synthesis of flavonoids. Our results also provide a valuable resource for further study on safflower.

SUBMITTER: Chen J

PROVIDER: S-EPMC6057038 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Full-length transcriptome sequences and the identification of putative genes for flavonoid biosynthesis in safflower.

Chen Jiang J Tang Xiaohui X Ren Chaoxiang C Wei Bin B Wu Yiyun Y Wu Qinghua Q Pei Jin J

BMC genomics 20180724 1

<h4>Background</h4>The flower of the safflower (Carthamus tinctorius L.) has been widely used in traditional Chinese medicine for the ability to improve cerebral blood flow. Flavonoids are the primary bioactive components in safflower, and their biosynthesis has attracted widespread interest. Previous studies mostly used second-generation sequencing platforms to survey the putative flavonoid biosynthesis genes. For a better understanding of transcription data and the putative genes involved in f ...[more]

PMID: 30041604

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Project description:BackgroundSafflower (Carthamus tinctorius L.) is a domesticated species with a long history of cultivation and widespread distribution across the globe, and light plays an important role in controlling its distribution boundary. Flowers from safflower have been widely used in traditional Chinese medicine because of their ability to improve cerebral blood flow. Flavonoids are the main active compounds in safflower and have many pharmacological effects. In this study, we aimed to explore the relationship between different light intensities and flavonoid biosynthesis in safflower flowers cultivated in greenhouse.MethodsThe transcriptome of safflower flowers grown under different light intensities were sequenced through BGISEQ-500 platform. After assembled and filtered, Unigenes were annotated by aligning with seven functional databases. Differential expression analysis of two samples was performed with the DEseq2 package. Differentially expressed genes (DEGs) related with flavonoids biosynthesis were analyzed by Real-time PCR (RT-PCR). Flavonoids accumulation in flowers were determined by high performance liquid chromatography and spectrophotometer.ResultsTranscriptome analysis of safflower flowers cultivated under different light intensities was performed. A total of 99.16 Gb data were obtained, and 78,179 Unigenes were annotated. Among the DEGs, 13 genes were related to flavonoid biosynthesis. The differential expressions of seven key genes were confirmed by RT-PCR. In addition, the levels of some flavonoids were measured in safflower flowers grown under different light intensities. CtHCT3 gene expression showed a significantly negative correlation with kaempferol content in safflower grown under different light intensities.ConclusionOur results strongly suggested that the reduction in light intensity in a suitable range promoted flavonoid biosynthesis in safflower flowers. We suggest that the expressions of HCT genes played an important role in flavonoid accumulation in safflower flowers. Our study lays a foundation for further research on the effects of light on flavonoid biosynthesis in safflower.

Project description:BackgroundSafflower (Carthamus tinctorius L.) is an important cash crop, of which the dried tube flower is not only an important raw material for dyes and cosmetics but also an important herb widely used in traditional Chinese medicine. The pigment and bioactive compounds are composed of flavonoids (mainly quinone chalcones), and studies have reported that MeJA can promote the biosynthesis of quinone chalcones, but the mechanism underlying the effect of MeJA in safflower remains unclear. Here, we attempt to use metabolomics and transcriptome technologies to analyse the molecular mechanism of flavonoid biosynthesis under MeJA treatment in safflower.ResultsBased on a UHPLC-ESI-MS/MS detection platform and a self-built database (including hydroxysafflor yellow A, HSYA), a total of 209 flavonoid metabolites were detected, and 35 metabolites were significantly different after treatment with MeJA. Among them, 24 metabolites were upregulated upon MeJA treatment, especially HSYA. Eleven metabolites were downregulated after MeJA treatment. Integrated metabolomics and transcriptome analysis showed that MeJA might upregulate the expression of upstream genes in the flavonoid biosynthesis pathway (such as CHSs, CHIs and HCTs) and downregulate the expression of downstream genes (such as F3Ms, ANRs and ANSs), thus promoting the biosynthesis of quinone chalcones, such as HSYA. The transcription expressions of these genes were validated by real-time PCR. In addition, the promoters of two genes (CtCHI and CtHCT) that were significantly upregulated under MeJA treatment were cloned and analysed. 7 and 3 MeJA response elements were found in the promoters, respectively.ConclusionsMeJA might upregulate the expression of the upstream genes in the flavonoid biosynthesis pathway and downregulate the expression of the downstream genes, thus promoting the biosynthesis of quinone chalcones. Our results provide insights and basic data for the molecular mechanism analysis of flavonoid synthesis in safflower under MeJA treatment.

Project description:Artemisia argyi is an important medicinal plant widely utilized for moxibustion heat therapy in China. The terpenoid biosynthesis process in A. argyi is speculated to play a key role in conferring its medicinal value. However, the molecular mechanism underlying terpenoid biosynthesis remains unclear, in part because the reference genome of A. argyi is unavailable. Moreover, the full-length transcriptome of A. argyi has not yet been sequenced. Therefore, in this study, de novo transcriptome sequencing of A. argyi's root, stem, and leaf tissues was performed to obtain those candidate genes related to terpenoid biosynthesis, by combining the PacBio single-molecule real-time (SMRT) and Illumina sequencing NGS platforms. And more than 55.4 Gb of sequencing data and 108,846 full-length reads (non-chimeric) were generated by the Illumina and PacBio platform, respectively. Then, 53,043 consensus isoforms were clustered and used to represent 36,820 non-redundant transcripts, of which 34,839 (94.62%) were annotated in public databases. In the comparison sets of leaves vs roots, and leaves vs stems, 13,850 (7,566 up-regulated, 6,284 down-regulated) and 9,502 (5,284 up-regulated, 4,218 down-regulated) differentially expressed transcripts (DETs) were obtained, respectively. Specifically, the expression profile and KEGG functional enrichment analysis of these DETs indicated that they were significantly enriched in the biosynthesis of amino acids, carotenoids, diterpenoids and flavonoids, as well as the metabolism processes of glycine, serine and threonine. Moreover, multiple genes encoding significant enzymes or transcription factors related to diterpenoid biosynthesis were highly expressed in the A. argyi leaves. Additionally, several transcription factor families, such as RLK-Pelle_LRR-L-1 and RLK-Pelle_DLSV, were also identified. In conclusion, this study offers a valuable resource for transcriptome information, and provides a functional genomic foundation for further research on molecular mechanisms underlying the medicinal use of A. argyi leaves.

Project description:The seed is the pharmaceutical and breeding organ of Cassia obtusifolia, a well-known medical herb containing aurantio-obtusin (a kind of anthraquinone), food, and landscape. In order to understand the molecular mechanism of the biosynthesis of aurantio-obtusin, seed formation and development, and stress response of C. obtusifolia, it is necessary to understand the genomics information. Although previous seed transcriptome of C. obtusifolia has been carried out by short-read next-generation sequencing (NGS) technology, the vast majority of the resulting unigenes did not represent full-length cDNA sequences and supply enough gene expression profile information of the various organs or tissues. In this study, fifteen cDNA libraries, which were constructed from the seed, root, stem, leaf, and flower (three repetitions with each organ) of C. obtusifolia, were sequenced using hybrid approach combining single-molecule real-time (SMRT) and NGS platform. More than 4,315,774 long reads with 9.66 Gb sequencing data and 361,427,021 short reads with 108.13 Gb sequencing data were generated by SMRT and NGS platform, respectively. 67,222 consensus isoforms were clustered from the reads and 81.73% (61,016) of which were longer than 1000 bp. Furthermore, the 67,222 consensus isoforms represented 58,106 nonredundant transcripts, 98.25% (57,092) of which were annotated and 25,573 of which were assigned to specific metabolic pathways by KEGG. CoDXS and CoDXR genes were directly used for functional characterization to validate the accuracy of sequences obtained from transcriptome. A total of 658 seed-specific transcripts indicated their special roles in physiological processes in seed. Analysis of transcripts which were involved in the early stage of anthraquinone biosynthesis suggested that the aurantio-obtusin in C. obtusifolia was mainly generated from isochorismate and Mevalonate/methylerythritol phosphate (MVA/MEP) pathway, and three reactions catalyzed by Menaquinone-specific isochorismate synthase (ICS), 1-deoxy-d-xylulose-5-phosphate synthase (DXS) and isopentenyl diphosphate (IPPS) might be the limited steps. Several seed-specific CYPs, SAM-dependent methyltransferase, and UDP-glycosyltransferase (UDPG) supplied promising candidate genes in the late stage of anthraquinone biosynthesis. In addition, four seed-specific transcriptional factors including three MYB Transcription Factor (MYB) and one MADS-box Transcription Factor (MADS) transcriptional factors) and alternative splicing might be involved with seed formation and development. Meanwhile, most members of Hsp20 genes showed high expression level in seed and flower; seven of which might have chaperon activities under various abiotic stresses. Finally, the expressional patterns of genes with particular interests showed similar trends in both transcriptome assay and qRT-PCR. In conclusion, this is the first full-length transcriptome sequencing reported in Caesalpiniaceae family, and thus providing a more complete insight into aurantio-obtusin biosynthesis, seed formation and development, and stress response as well in C. obtusifolia.

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Full-length transcriptome sequences and the identification of putative genes for flavonoid biosynthesis in safflower.

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Full-length transcriptome sequences and the identification of putative genes for flavonoid biosynthesis in safflower.

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