Public Health · Vol. 10, No. 2 · 2026 · pp. 1–9 · e-ISSN 2472-3878

Effects of Temperature and Centrifugation Conditions on DNA Isolation From Different Leaf Ages In Aconitum Heterophyllum Wall. Ex Royle

M. Sainath Rao, Satyam Kumar Mehta, Anjali, M. Ramanjineyulu, Vijay Kant Purohit, Jaidev Chauhan

DOI: https://doi.org/10.17140/phoj.10.02.1 · Published 2026-04-22 · Licence: https://creativecommons.org/licenses/by/4.0 · © 2026 The Author(s)

Also available as PDF (figures and tables appear in the PDF).

Abstract

Aconitum heterophyllum Wall. ex Royle, commonly known as Atees, is a critically endangered medicinal herb endemic to the Northwestern Himalayan region, valued for its therapeutic alkaloid atisine. The extraction of high-quality genomic DNA from this species presents significant challenges due to elevated concentrations of polysaccharides, polyphenolic compounds and secondary metabolites inherent to high-altitude medicinal plants. This study systematically evaluated the effects of leaf developmental stage (young, middle-aged and mature) and centrifugation conditions on DNA yield and purity using a modified CTAB protocol. Samples were collected from Ghesh village, Chamoli district, Uttarakhand and DNA was extracted under varying parameters: 10,000 rpm at 25°C, 12,000 rpm at 30°C and 15,000 rpm at 35°C. Spectrophotometric and electrophoretic analyses revealed that young leaves produced the highest DNA yield (847 µg/mL) and optimal purity ratios (A₂₆₀/A₂₈₀ = 1.82, A₂₆₀/A₂₃₀ = 2.15) under 10,000 rpm at 25°C, whereas higher centrifugation speeds and temperatures resulted in reduced quality. Optimal DNA extraction was achieved at 10,000 rpm and 25°C, demonstrating that elevated centrifugation speeds and temperatures inversely correlate with DNA quality. The inclusion of 1% PVP and 0.2% β-mercaptoethanol in the CTAB buffer effectively mitigated interference from secondary metabolites. RAPD-PCR amplification using primer OPA-1 (CAGGCCCTTC) generated clear and distinct banding patterns (7-9 bands) from optimally extracted DNA, confirming its suitability for molecular applications. These findings establish a reproducible, cost-effective protocol for high-quality genomic DNA extraction from A. heterophyllum , providing methodological guidance for molecular characterization, genetic diversity assessment and conservation strategies for endangered Himalayan medicinal flora.

Keywords: Aconitum heterophyllum; CTAB extraction; centrifugation optimisation; leaf age variation; DNA purity; RAPD markers; medicinal plants; Himalayan biodiversity; conservation genetic; Aconitumheterophyllum; CTABextraction; centrifugationoptimisation; leafagevariation; RAPDmarkers; medicinalplants; Himalayanbiodiversity; conservationgenetic

Acknowledgement

Theauthorswishtoexpresstheirsinceregratitudeto

Dr. VijayKantPurohitforhisinvaluablesupervision etal

andtoDirectorMohanChandraNautiyalforhiscontin-

uoussupport.WearethankfultoAssistantProfessors A.heterophyllum

DrPrattiPrasad,Dr.VijayLaxmiTrivediandDr.Babita

Patnifortheirvaluableguidance.Wealsoacknowledge

researchscholarseniorJaidevChauhan, postgraduetal

ateseniorAnshikaPokhriyal,andjuniorGekenRiba,

ApekshaNegifortheircooperationandassistance

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