Abstract:
Background: Ajuga bracteosais a prominent source of therapeutically active compounds. However, the yield of these phytochemicals is extremely low in wild A. bracteosa which can be enhanced by somatic embryogenesis as well as metabolic engineering techniques. Thus, present study aimed to develop an efficient protocol for high frequency somatic embryogenesis and optimization of the conditions for the stable genetic transformation of A. bracteosa with root oncogenic loci (rol)A and C genes. Moreover, the possible impact of rolgenes transformation on phytochemistry and pharmacological properties (antioxidant, antibacterial, antiproliferative, and antidiabetic) was also evaluated. Methods: For somatic embryogenesis, various combinations of auxins and cytokinin's were employed on tissue cultured explants. Additionally, Agrobacterium tumefaciens strain GV3101 harboring the rolA, rolC and nptII genes was used for transformation of leaf discs and nodal regions of in vitro grown A. bracteosa. Successful integration of rol genes in putative transformants and their copy number was confirmed by conventional PCR, RT-PCR and southern blot techniques, respectively. Then the plant extracts were prepared to check the phenolic and flavonoid content, antioxidant, and antioxidant activity. Whereas, cytotoxic activity against MCF-7 human breast cancer cell line, and antidiabetic activity via DPP-4 assay were also assessed. Besides, the plant metabolites were quantified by ESI-QTOF-MS analysis. Among various explants, leaves showed maximum response for callus induction with the hormonal combination of 1mg/L NAA + 0.5 mg/L BAP. Positive PCR results were obtained for rolA (308 bp), rolC (540 bp) and nptII(780 bp) genes in gDNA of transformed plants. RT-PCR indicated the varied expression of rolA and rolC transcripts in different transgenic lines. A4 line showed the high transcript level compared to rest of rolA transgenic lines. While among rolC transgenic lines, C1 had maximum transcript level. Southern blot analysis of PCR-positive plants showed that one copy of bacterial gene was inserted in all the transformants except A4 and C1 having two copies. These 2 lines (A4 and C1) showed maximum phenolic and flavonoid content, antioxidant potential, antibacterial activity, antiproliferative, and antidiabetic activities when compared with other transgenic and wild type plants. These 2 lines also had the highest number of primary and secondary metabolites. This study opens up the avenues for effective means of preserving and multiplying this endangered medicinal plant to cope with its overexploitation. Agrobacterium rol A and C can account for the increased level of phytochemicals and significant biological activity of the transgenic lines. Further studies that can be carried out to recognize genes involved in biosynthesis of key metabolites identified, which can be used in metabolic engineering for obtaining high yield of these pharmaceutically important metabolites to meet market demands.
Page(s):
59-59
DOI:
DOI not available
Published:
Journal: Abstract Book on International Conference on Food and Applied Sciences (ICFAS-23) 3-5 August 23, Volume: 0, Issue: 0, Year: 2023
Keywords:
antidiabetic activity
,
Antiproliferative activity
,
Somatic embryogenesis
,
Ajuga bracteosa
,
LCMS