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A survey of transcriptome complexity using PacBio single-molecule real-time analysis combined with Illumina RNA sequencing for a better understanding of ricinoleic acid biosynthesis in Ricinus communis.

ABSTRACT: BACKGROUND:Ricinus communis is a highly economically valuable oil crop plant from the spurge family, Euphorbiaceae. However, the available reference genomes are incomplete and to date studies on ricinoleic acid biosynthesis at the transcriptional level are limited. RESULTS:In this study, we combined PacBio single-molecule long read isoform and Illumina RNA sequencing to identify the alternative splicing (AS) events, novel isoforms, fusion genes, long non-coding RNAs (lncRNAs) and alternative polyadenylation (APA) sites to unveil the transcriptomic complexity of castor beans and identify critical genes related to ricinoleic acid biosynthesis. Here, we identified 11,285 AS-variants distributed in 21,448 novel genes and detected 520 fusion genes, 320 lncRNAs and 9511 (APA-sites). Furthermore, a total of 6067, 5983 and 4058 differentially expressed genes between developing beans of the R. communis lines 349 and 1115 with extremely different oil content were identified at 7, 14 and 21?days after flowering, respectively. Specifically, 14, 18 and 11 DEGs were annotated encoding key enzymes related to ricinoleic acid biosynthesis reflecting the higher castor oil content of 1115 compared than 349. Quantitative real-time RT-PCR further validated fifteen of these DEGs at three-time points. CONCLUSION:Our results significantly improved the existed gene models of R. communis, and a putative model of key genes was built to show the differences between strains 349 and 1115, illustrating the molecular mechanism of castor oil biosynthesis. A multi-transcriptome database and candidate genes were provided to further improve the level of ricinoleic acid in transgenic crops.

SUBMITTER: Wang L

PROVIDER: S-EPMC6555039 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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A survey of transcriptome complexity using PacBio single-molecule real-time analysis combined with Illumina RNA sequencing for a better understanding of ricinoleic acid biosynthesis in Ricinus communis.

Wang Lijun L Jiang Xiaoling X Wang Lei L Wang Wei W Fu Chunling C Yan Xingchu X Geng Xinxin X

BMC genomics 20190606 1

<h4>Background</h4>Ricinus communis is a highly economically valuable oil crop plant from the spurge family, Euphorbiaceae. However, the available reference genomes are incomplete and to date studies on ricinoleic acid biosynthesis at the transcriptional level are limited.<h4>Results</h4>In this study, we combined PacBio single-molecule long read isoform and Illumina RNA sequencing to identify the alternative splicing (AS) events, novel isoforms, fusion genes, long non-coding RNAs (lncRNAs) and ...[more]

PMID: 31170917

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Project description:Microsomal membrane preparations from the developing endosperm of castor bean (Ricinus communis) catalysed the transfer of oleate from [14C]oleoyl-CoA to phosphatidylcholine (PtdCho). In the presence of NADH, radioactive ricinoleate (12-hydroxyoctadec-9-enoate) was synthesized from [14C]oleate, and this was largely recovered in PtdCho and as free fatty acid. The addition of unlabelled ricinoleoyl-CoA to these incubation mixtures did not increase the low [14C]ricinoleate concentration found in the acyl-CoA fraction nor decrease the [14C]ricinoleate concentration in PtdCho and free fatty acid, and thus no evidence was obtained for a hydroxylation with oleoyl-CoA as a substrate. The addition of NADH, necessary for the formation of ricinoleate, caused a decrease of the total radioactivity in PtdCho with a corresponding increase in the amount of label in free ricinoleic acid. This increase was due to the action of a phospholipase A, which released ricinoleic acid but not oleic acid from PtdCho. Such a phospholipase activity, attacking ricinoleoyl-PtdCho but not oleoyl-PtdCho, was also demonstrated in microsomal preparations from developing cotyledons of safflower and oil-seed rape. An analysis of the acyl groups at different positions in microsomal PtdCho of castor bean showed that ricinoleate was almost entirely associated with position sn-2. Likewise the [14C]ricinoleate in [14C]PtdCho formed after incubations with microsomal preparations with NADH and [14C]oleoyl-CoA resided in position sn-2 with none in position sn-1. In contrast, the [14C]linoleate formed by desaturation of [14C]oleoyl-PtdCho was present at both positions. In the presence of ATP, CoA and Mg2+, the ricinoleate acid released from PtdCho was activated to ricinoleoyl-CoA. The ricinoleoyl-CoA was an efficient acyl donor in the acylation of glycerol 3-phosphate (Gro3P) to yield phosphatidic acid and triacylglycerols. In microsomal preparations incubated with an equimolar mixture of [14C]oleoyl-CoA and [14C]ricinoleoyl-CoA in the presence of Gro3P, only a minor amount of [14C]ricinoleate entered PtdCho, and this was believed to be via the exchange of phosphocholine groups between a diacylglycerol pool and the PtdCho. On the basis of our results, a scheme of ricinoleate formation and its incorporation into triacylglycerols in castor-bean endosperm is proposed.

Dataset Information

A survey of transcriptome complexity using PacBio single-molecule real-time analysis combined with Illumina RNA sequencing for a better understanding of ricinoleic acid biosynthesis in Ricinus communis.

Publications

A survey of transcriptome complexity using PacBio single-molecule real-time analysis combined with Illumina RNA sequencing for a better understanding of ricinoleic acid biosynthesis in Ricinus communis.

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