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Transcriptome Analysis and Identification of Genes Related to Biosynthesis of Anthocyanins and Kaempferitrin in Kenaf (Hibiscus cannabinus L.)

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Abstract

Kenaf is a valuable fiber and medicinal crop. Here, we performed RNA-sequencing to obtain comprehensive transcriptome information and identify putative genes involved in biosynthetic pathways of anthocyanins and kaempferitrin in kenaf. A total of 39.6 Gb reads were generated for six kenaf accessions. Individual de novo assembly of each accession and clustering of whole transcriptome contigs finally yielded 299,880 representative transcripts having an average length of 1217 bp, of which 231,825 (77.3%) were annotated against various databases. Expression profiling of the transcripts identified 6,592 differentially expressed genes (DEGs) among the three accessions, C-14 (a local landrace) and two of its mutant varieties, Jeokbong (purple leaves and stems) and Baekma (light green leaves and stems). In addition to their altered coloration, leaves of these two mutants had significantly different anthocyanin and kaempferitrin contents. We constructed anthocyanin and kaempferitrin biosynthetic pathways and identified 671 transcripts mapping to the entire pathway. In addition, 29 DEGs assigned to eight structural genes and 41 DEGs related to three transcription factor families were identified in high anthocyanin- and kaempferitrin-accumulating mutant Jeokbong. Our results provide a large transcriptome library pool for understanding gene functions and should also be useful in further studies of flavonoid biosynthesis in kenaf.

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Acknowledgements

This work was supported by the research program of Korea Atomic Energy Research Institute, Republic of Korea.

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  1. Jae Il Lyu and Hong-Il Choi contributed equally to the manuscript.

Authors and Affiliations

  1. Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup, 56212, Republic of Korea

    Jae Il Lyu, Hong-Il Choi, Jaihyunk Ryu, Soon-Jae Kwon, Yeong Deuk Jo, Min Jeong Hong, Jin-Baek Kim, Joon-Woo Ahn & Si-Yong Kang

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Contributions

HIC, JR, SJK, and SYK conceived and designed the experiments. JIL, HIC, and JR performed experiments and transcriptome analysis. JIL, HIC, YDJ, MJH, JBK, and JWA analyzed and interpreted the experimental data. JIL and HIC wrote the manuscript. All authors reviewed the results and approved the final version of the manuscript.

Corresponding author

Correspondence to Hong-Il Choi.

Electronic supplementary material

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12374_2020_9227_MOESM1_ESM.tif

Supplementary file1 Fig. S1. Comparison of morphological characteristics of C-14, Baekma (BM), and Jeokbong (JB). (A) Leaf. (B) Stem. (C) Whole plants in field. (TIF 21119 kb)

12374_2020_9227_MOESM2_ESM.tif

Supplementary file2 Fig. S2. A Venn diagram of differentially expressed genes (DEGs) involved in anthocyanin and kaempferitrin biosynthetic pathways in kenaf. In the case of enzymes with more than one identified DEG, the number of DEGs is given in parentheses. (TIF 3708 kb)

12374_2020_9227_MOESM3_ESM.tif

Supplementary file3 Fig. S3. A Venn diagram of differentially expressed genes containing transcription factor (TF) domains regulating flavonoid biosynthetic pathway genes in kenaf. (TIF 3836 kb)

12374_2020_9227_MOESM4_ESM.tif

Supplementary file4 Fig. S4. Validation of differentially expressed genes related to anthocyanin and kaempferitrin biosynthesis by qRT-PCR. (A–L) Transcript expression levels for 4CL involved in phenylpropanoid biosynthesis (A–B), CHS, CHI, F3H, DFR, ANS, and FLS involved in flavonoid biosynthesis (C–J), and 3GT involved anthocyanin biosynthesis (K–L). Vertical bars are relative expression values (left y-axis) normalized to the abundance of the reference genes (HcACT7 for left graphs and HcPP2A for right graphs) by qRT-PCR, and lines indicate expression levels calculated from FPKM values (right y-axis) obtained by RNA sequencing. qRT-PCR data are based on three biological replicates and represent the mean ± SE of triplicate repeats. (TIF 10227 kb)

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Lyu, J.I., Choi, HI., Ryu, J. et al. Transcriptome Analysis and Identification of Genes Related to Biosynthesis of Anthocyanins and Kaempferitrin in Kenaf (Hibiscus cannabinus L.). J. Plant Biol. 63, 51–62 (2020). https://doi.org/10.1007/s12374-020-09227-9

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