Abstract
Isatis indigotica Fort. is a popular herb in traditional Chinese medicine. It possesses pharmacological activities against various diseases, particularly leukemia. Indigoid alkaloids are its main bioactive ingredients. A relatively low nitrogen supply increases indigoid content in I. indigotica leaves, yet the mechanisms that regulate indigoid biosynthesis are unknown. To uncover regulatory mechanisms, we performed transcriptome sequencing of leaf samples from I. indigotica exposed to deficient (0 mM), low (7.5 mM), and high (15 mM) concentrations of NO3−. We generated 38,990 unigenes with an N50 value of 1923 bp, of which 33,007 unigenes (84.65%) were annotated in at least one database. In comparison with deficient N, we identified 6911 and 4845 differentially expressed genes (DEGs) in high N and low N. Cluster analysis of these DEGs showed a unique expression pattern in the N-supplied compared with the N-free condition. Twenty-one genes related to nitrogen uptake, transport, and assimilation were highly expressed in N-deficient conditions. Further, 63 putative genes that encode for enzymes involved in indigoid biosynthesis in I. indigotica leaf were identified and analyzed. Thirteen genes involved in indole modification were expressed more highly in low N; their high expression level may explain the increase in indigoid alkaloids in low N. Notably, phylogenetic analysis showed that IiTSA3 may relate to genes involved in indole heterocycle biosynthesis. Additionally, 491 differentially expressed TFs were also identified. Our findings will enrich gene resources for elucidating the molecular mechanisms of indigoid alkaloid biosynthesis in I. indigotica leaves under different nitrogen supplies.
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The authors are grateful to the Institute of Food Crops, Jiangsu Academy Agriculture sciences, China for providing lab facilities and experimental support. The authors thank AiMi Academic Services (www.aimieditor.com) for English language editing and review services.
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This research was funded by a Special subsidy for TCM public health services in 2018 “National traditional Chinese medicine resources survey project” (caise [2018] No.43), National Natural Science Foundation of China (No. 31171486), and Zhenjiang Jinshan Talents in Jiangsu Province of China Modern Agricultural Leaders (Innovation) Project (2018).
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XT and RQ conceived and designed the experiments. LS, YC, HZ, YM, and RQ performed the RNA-Seq and data analysis; RQ and YC performed qRT-PCR measurement and analysis. RQ and YM wrote the paper.
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Supplementary Materials: Supplementary materials are at XXX. We have deposited the raw data in in the NCBI's Sequence Read Archive (accession SRP201538), and the 'Transcriptome Shotgun Assembly' project has been deposited at DDBJ/EMBL/GenBank (accession GHNT01000000.) Supplementary file1 (DOCX 17 kb)
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Qu, R., Cao, Y., Zhang, H. et al. Transcriptome Analysis of Nitrogen Metabolism, Transcription Factors, and Indigoid Biosynthesis in Isatis indigotica Fort. Response to Nitrogen Availability. J Plant Growth Regul 40, 1181–1197 (2021). https://doi.org/10.1007/s00344-020-10178-1
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DOI: https://doi.org/10.1007/s00344-020-10178-1