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A Miniprep DNA Verifiable Method from Denatured Samples for DNA Barcoding and Sequencing Applications

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Proceedings of the National Academy of Sciences, India Section B: Biological Sciences Aims and scope Submit manuscript

Abstract

Exploration on DNA-based investigations is frequently obscured by being constrained to work in suboptimal conditions that incorporate minimal sampling regimes, crucial tissue type, and amount. As nucleic acids are the source to explain different inquiries in DNA-involved investigations there is a need for a trendy strategy to affirm entire genome from complicated heterogeneous vegetation. In this verification, authors modified CTAB DNA extracting method to develop a fast and precise protocol for dehydrated biological material and extremely processed commercial samples. The tea plant, Camellia sinensis, has the importation of barely recognizable raw product when fermented and high adulteration risk commercially is the advantage noted to study the plant. To reaffirm the success of the present problem the DNA extracted from the protocol is subjected to molecular manipulations like sequencing of DNA with CBOL recommended plant barcoding markers like rbcL, matK, trnH-psbA, rpoC, rpoB, ycf1, ITS2 and created a standard reference library for south Indian tea clones to authenticate unknown taxa. This protocol was standardized with the initial screening of ten south Indian tea clones and five commercial samples. After successful trials, the modified CTAB manual was applied to 40 dehydrated tea cultivars and ten commercially available black dust and green tea. Authors have also verified the tea cultivars sequences extracted by this method are applicable to generate DNA barcodes. The tested method's success in extremely processed specimens proves the modified CTAB method as a promising alternative among all reported CTAB methods and costly isolation kits for incomplete specimens.

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Acknowledgements

The authors thank the National Tea Research Foundation (NTRF), Government of India, (S.O: NTRF: 190/2016) for the financial support. The authors also thank the Karunya Institute of Technology and Sciences for providing the research facilities.

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Authors and Affiliations

  1. Algae Biomass Research Laboratory, Department of Biotechnology, Karunya Institute of Technology and Sciences (Deemed to be University), Coimbatore, Tamil Nadu, 641114, India

    Steffi Avarave & Jibu Thomas

Authors
  1. Steffi Avarave
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  2. Jibu Thomas
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Contributions

Steffi Avarave conceived the research and designed the experiments. Steffi Avarave performed the experiments, analyzed and interpreted the data. Steffi Avarave wrote the original draft. Jibu Thomas supervised and edited the manuscript. All authors edited and approved the final manuscript.

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Correspondence to Jibu Thomas.

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Significance statement The checked protocol ensures getting dependable and explicit distinguishing proof using DNA barcoding, which could magnify the information gleaned from the plant chloroplast, nuclear markers and even adulterant identification in commercial tea dust.

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Avarave, S., Thomas, J. A Miniprep DNA Verifiable Method from Denatured Samples for DNA Barcoding and Sequencing Applications. Proc. Natl. Acad. Sci., India, Sect. B Biol. Sci. 92, 329–340 (2022). https://doi.org/10.1007/s40011-021-01334-y

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  • DOI: https://doi.org/10.1007/s40011-021-01334-y

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