Molecular cloning and In-silico characterization of NAC86 of Kalmegh (Andrographis paniculata)

Kumar, Ramesh; Kumar, Chavlesh; Jain, Rishu; Maurya, Avantika; Kumar, Ashok; Kumari, Abha; Singh, Rakesh K.

doi:10.5958/0974-0112.2022.00002.0

Cited by 2 publications

(5 citation statements)

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“…The WRKY TF are among the largest families of transcriptional regulators mainly involved in plant growth and developmental processes [61] and the NAC family of TFs is involved in the regulation of biotic and abiotic stress responses [62]. The PCR amplification of TF gene-specific primers from three important TF families, i.e., bHLH, MYB, and NAC, in two different A. paniculata accessions, as well as molecular cloning and characterization of the NAC86 gene by Ramesh et al [63], showed the quality of our transcriptome assembly. A total of 106 transcripts have been identified as being related to the cytochrome p450 enzyme group, and 766 transcripts are related to protein kinases; a similar type of transcript has already been reported in T. cordifolia [64], an important medicinal plant that also produces secondary metabolites using terpenoid pathway.…”

Section: Discussionmentioning

confidence: 99%

“…For any plant species, the extent of available diversity in their germplasm is an invaluable asset for their appropriate utilization. Kumar et al [63] performed cloning and characterization of the full-length NAC86 TF gene associated with the secondary metabolite biosynthetic pathway using the ApTransDB database. This study shows the possibilities and the utility of this database, which can be further extended for mining transcripts/genes associated with other related pathways.…”

Section: Discussionmentioning

confidence: 99%

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De Novo Transcriptome Profiling for the Generation and Validation of Microsatellite Markers, Transcription Factors, and Database Development for Andrographis paniculata

Singh

Mahato

et al. 2023

IJMS

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Andrographis paniculata belongs to the family Acanthaceae and is known for its medicinal properties owing to the presence of unique constituents belonging to the lactones, diterpenoids, diterpene glycosides, flavonoids, and flavonoid glycosides groups of chemicals. Andrographolide, a major therapeutic constituent of A. paniculata, is extracted primarily from the leaves of this plant and exhibits antimicrobial and anti-inflammatory activities. Using 454 GS-FLX pyrosequencing, we have generated a whole transcriptome profile of entire leaves of A. paniculata. A total of 22,402 high-quality transcripts were generated, with an average transcript length and N50 of 884 bp and 1007 bp, respectively. Functional annotation revealed that 19,264 (86%) of the total transcripts showed significant similarity with the NCBI-Nr database and were successfully annotated. Out of the 19,264 BLAST hits, 17,623 transcripts were assigned GO terms and distributed into three major functional categories: molecular function (44.62%), biological processes (29.19%), and cellular component (26.18%) based on BLAST2GO. Transcription factor analysis showed 6669 transcripts, belonging to 57 different transcription factor families. Fifteen TF genes that belong to the NAC, MYB, and bHLH TF categories were validated by RT PCR amplification. In silico analysis of gene families involved in the synthesis of biochemical compounds having medicinal values, such as cytochrome p450, protein kinases, heat shock proteins, and transporters, was completed and a total of 102 different transcripts encoding enzymes involved in the biosynthesis of terpenoids were predicted. Out of these, 33 transcripts belonged to terpenoid backbone biosynthesis. This study also identified 4254 EST-SSRs from 3661 transcripts, representing 16.34% of the total transcripts. Fifty-three novel EST-SSR markers generated from our EST dataset were used to assess the genetic diversity among eighteen A. paniculata accessions. The genetic diversity analysis revealed two distinct sub-clusters and all accessions based on the genetic similarity index were distinct from each other. A database based on EST transcripts, EST-SSR markers, and transcription factors has been developed using data generated from the present study combined with available transcriptomic resources from a public database using Meta transcriptome analysis to make genomic resources available in one place to the researchers working on this medicinal plant.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

De Novo Transcriptome Profiling for the Generation and Validation of Microsatellite Markers, Transcription Factors, and Database Development for Andrographis paniculata

Singh

Mahato

et al. 2023

IJMS

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“…Three replicates of each qPCR reaction were performed, and a non-template negative control (NTC) was also performed. The relative expression level was calculated using the 2 −∆∆CT method and considering the actin gene of A. paniculata for normalization [55,72]. During the expression analysis under the MeJA and ABA treatments, all the expressions were compared with the control (untreated).…”

Section: Expression Analysis Using Real-time Quantitative Pcr (Qpcr)mentioning

confidence: 99%

“…During the expression analysis under the MeJA and ABA treatments, all the expressions were compared with the control (untreated). The housekeeping gene actin was used as an endogenous control during expression analysis [55]. Based on the literature search, the andrographolide biosynthesis pathways, such as MVA (mevalonate pathway) and MEP (2-C-methyl-D-erythritol 4-phosphate) pathways, were examined.…”

Section: Expression Analysis Using Real-time Quantitative Pcr (Qpcr)mentioning

confidence: 99%

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Isolation, Characterization, and Expression Analysis of NAC Transcription Factor from Andrographis paniculata (Burm. f.) Nees and Their Role in Andrographolide Production

Kumar,

Roy Choudhury

et al. 2024

Genes

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Andrographis paniculata (Burm. f.) Nees is an important medicinal plant known for its bioactive compound andrographolide. NAC transcription factors (NAM, ATAF1/2, and CUC2) play a crucial role in secondary metabolite production, stress responses, and plant development through hormonal signaling. In this study, a putative partial transcript of three NAC family genes (ApNAC83, ApNAC21 22 and ApNAC02) was used to isolate full length genes using RACE. Bioinformatics analyses such as protein structure prediction, cis-acting regulatory elements, and gene ontology analysis were performed. Based on in silico predictions, the diterpenoid profiling of the plant’s leaves (five-week-old) and the real-time PCR-based expression analysis of isolated NAC genes under abscisic acid (ABA) treatment were performed. Additionally, the expression analysis of isolated NAC genes under MeJA treatment and transient expression in Nicotiana tabacum was performed. Full-length sequences of three members of the NAC transcription factor family, ApNAC83 (1102 bp), ApNAC21 22 (996 bp), and ApNAC02 (1011 bp), were isolated and subjected to the promoter and gene ontology analysis, which indicated their role in transcriptional regulation, DNA binding, ABA-activated signaling, and stress management. It was observed that ABA treatment leads to a higher accumulation of andrographolide and 14-deoxyandrographolide content, along with the upregulation of ApNAC02 (9.6-fold) and the downregulation of ApNAC83 and ApNAC21 22 in the leaves. With methyl jasmonate treatment, ApNAC21 22 expression decreased, while ApNAC02 increased (1.9-fold), with no significant change being observed in ApNAC83. The transient expression of the isolated NAC genes in a heterologous system (Nicotiana benthamiana) demonstrated their functional transcriptional activity, leading to the upregulation of the NtHMGR gene, which is related to the terpene pathway in tobacco. The expression analysis and heterologous expression of ApNAC21 22 and ApNAC02 indicated their role in andrographolide biosynthesis.

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Molecular cloning and In-silico characterization of NAC86 of Kalmegh (Andrographis paniculata)

Cited by 2 publications

References 0 publications

De Novo Transcriptome Profiling for the Generation and Validation of Microsatellite Markers, Transcription Factors, and Database Development for Andrographis paniculata

De Novo Transcriptome Profiling for the Generation and Validation of Microsatellite Markers, Transcription Factors, and Database Development for Andrographis paniculata

Isolation, Characterization, and Expression Analysis of NAC Transcription Factor from Andrographis paniculata (Burm. f.) Nees and Their Role in Andrographolide Production

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