Optimisation of DNA isolation and PCR techniques for beetle (Order: Coleoptera) specimens

Murthy, Meesala Krishna; Khandayataray, Pratima; Tara, Malsawmdawngzuali; Pori, Buragohain; Abinash, Giri; Gurusubramanian, Guruswami

doi:10.1007/s42690-022-00736-3

Cited by 1 publication

(1 citation statement)

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“…However, most of them are not cost effective, time consuming and labour intensive. Currently, the research purpose is to reduce operating time, labour and cost without compromising DNA quantity and quality too much [13]. A fast, simple and low cost extraction approach with sufficient yield and purity of DNA would greatly improve the overall efficiency of molecular analysis.…”

Section: Introductionmentioning

confidence: 99%

A Fast, Simple and Low-cost DNA Extraction Protocol from Common Ants and Beetles for Multiple Molecular Applications

Jiaying,

Junxia,

et al. 2024

BIO

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The rapid development of molecular biology tools in insect systematics, invasion research, evolutionary ecology and biodiversity analysis has led to faster and greater progress in understanding insect behavior and biology. Efficient DNA extraction is the foremost step and serves as the vital foundation. Several rapid DNA extraction methods have been established, which are often time-consuming and labour-intensive. Here, a simple, fast, low-cost DNA extraction protocol for common insect samples was developed basing on 28 specimens of 16 insect species (7 ants, 9 bark and ambrosia beetles). The new protocol was shown to be feasible and highly efficient by comparison with commercial kit in terms of DNA yield, purity and PCR sensitivity. The concentration of DNA through the new rapid method was higher than that through commercial kit, whether in ant or beetle samples. A better quality of DNA extracted via kit was indicated by A<sub>260</sub>/A<sub>280</sub> mostly ranging from 1.80 to 2.00. There was little difference between DNA extracted from adult and nymphal insects. PCR sensitivity of extracted DNA using both protocols was comparable. For nested PCR, amplification after two rounds yielded a bright signal using template DNA through both methods. But for PCR using primers of LCO1490 and HCO2198, the success ratio was lower (85.18%). Through BLAST, these amplicons were matched to related data with high identity. By combining this protocol with variable analysis platforms such as common PCR, loop-mediated isothermal amplification, and high throughput sequencing, it could assist insect diagnostics, biological surveys and invasion researches.

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Section: Introductionmentioning

confidence: 99%