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Endophytic Consortium With Diverse Gene-Regulating Capabilities of Benzylisoquinoline Alkaloids Biosynthetic Pathway Can Enhance Endogenous Morphine Biosynthesis in Papaver somniferum.

ABSTRACT: Secondary metabolite biosynthesis in medicinal plants is multi-step cascade known to be modulated by associated endophytes. While a single endophyte is not able to upregulate all biosynthetic steps, limiting maximum yield achievement. Therefore to compliment the deficient characteristics in an endophyte we tried consortium of endophytes to achieve maximum yield. Here, efforts were made to maximize the in planta morphine yield, using consortium of two endophytes; SM1B (Acinetobacter sp.) upregulating most of the genes of morphine biosynthesis except T6ODM and CODM, and SM3B (Marmoricola sp.) upregulating T6ODM and CODM in alkaloid-less Papaver somniferum cv. Sujata. Consortium-inoculation significantly increased morphine and thebaine content, and also increased the photosynthetic efficiency of poppy plants resulted in increased biomass, capsule weight, and seed yields compared to single-inoculation. The increment in morphine content was due to the modulation of metabolic-flow of key intermediates including reticuline and thebaine, via upregulating pertinent biosynthetic genes and enhanced expression of COR, key gene for morphine biosynthesis. This is the first report demonstrating the endophytic-consortium complimenting the functional deficiency of one endophyte by another for upregulating multiple genes of a metabolic pathway similar to transgenics (overexpressing multiple genes) for obtaining enhanced yield of pharmaceutically important metabolites.

SUBMITTER: Ray T

PROVIDER: S-EPMC6503101 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Endophytic Consortium With Diverse Gene-Regulating Capabilities of Benzylisoquinoline Alkaloids Biosynthetic Pathway Can Enhance Endogenous Morphine Biosynthesis in Papaver somniferum.

Ray Tania T Pandey Shiv S SS Pandey Alok A Srivastava Madhumita M Shanker Karuna K Kalra Alok A

Frontiers in microbiology 20190430

Secondary metabolite biosynthesis in medicinal plants is multi-step cascade known to be modulated by associated endophytes. While a single endophyte is not able to upregulate all biosynthetic steps, limiting maximum yield achievement. Therefore to compliment the deficient characteristics in an endophyte we tried consortium of endophytes to achieve maximum yield. Here, efforts were made to maximize the in planta morphine yield, using consortium of two endophytes; SM1B (Acinetobacter ...[more]

PMID: 31114562

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Project description:INTRODUCTION:Papaver somniferum is the commercial source of morphine and codeine. The isolation of effective genes involved in the morphine biosynthesis of P. somniferum is very important in the production of specific metabolites achieved using metabolic engi-neering techniques. In this pathway, the key enzyme COR is involved in the conversion of codeinone to codeine and morphinone to morphine. METHODS:the gene encoding of this enzyme was isolated using primers designed on the base of gene sequence available on (NCBI) for P. somniferum. This gene correct size around (960 bp) was first subcloned into pTZ57RIT vector then cloned into expression vectors (pBI121) between BamHI and SacI sites to allow the expression of cor gene driven by the cauliflower mosaic virus 35S pro-moter. The result was confirmed through different molecular methods e.g. PCR and en-zyme digestion by BamHI and SacI. The recombinant plasmid was transformed into the E. coli strain DH5? using a freeze-thaw method. Having selected positive colones on selection medium, plasmid was extracted by miniprep method and recombinant plasmids were selected based on PCR and digestion. The construct was then mobilized in Agrobacterium tumefaciens C58/pGV3850 (KmR RifR). After gene transformation to P. somniferum plants, the agroinfiltration method was also used for transient expression of COR enzyme. RESULTS:evaluation results showed that morphine and codeine were detectable in the leaves of transgenic plants containing cor transgene and there was significant difference in the final production. After completing this experiment for three times, results showed that in 11 sets from 15 sets of leaves experiment tested, main alkaloids (codeine, morphine, papaverin, noscapine and thebaine) were detectable. CONCLUSION:Whereas no signal was detected in non-infiltrated control leaves or in leaves infiltrated with non-recombinant bacteria for morphine and codeine, others such as thebaine and papaverine were detectable.

Dataset Information

Endophytic Consortium With Diverse Gene-Regulating Capabilities of Benzylisoquinoline Alkaloids Biosynthetic Pathway Can Enhance Endogenous Morphine Biosynthesis in Papaver somniferum.

Publications

Endophytic Consortium With Diverse Gene-Regulating Capabilities of Benzylisoquinoline Alkaloids Biosynthetic Pathway Can Enhance Endogenous Morphine Biosynthesis in <i>Papaver somniferum</i>.

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