Accurate determination of DNA yield from individual mosquitoes for population genomic applications

Wilding, Craig, Weetman, David ORCID: https://orcid.org/0000-0002-5820-1388, Steen, Keith ORCID: https://orcid.org/0000-0002-8933-8643 and Donnelly, Martin ORCID: https://orcid.org/0000-0001-5218-1497 (2009) 'Accurate determination of DNA yield from individual mosquitoes for population genomic applications'. Insect Science, Vol 16, Issue 4, pp. 361-363.

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Abstract

Accurate estimates of DNA quantity are likely to become increasingly important for successful genomic screening of insect populations via recently developed, highly multiplexed genotyping assays and high-throughput sequencing methods. Here we show that genomic DNA extractions from single Anopheles gambiae Giles using a standard commercial kit-based methodology yield extracts with concentrations below the linear range of spectrophotometric absorbance at 260 nm. Concentrations determined by spectrophotometry were not reproducible, and are therefore neither accurate nor reliable. However, DNA quantification using a fluorescent nucleic acid stain (PicoGreen((R))) gave highly reproducible concentration estimates, and indicated that, on average, single mosquitoes yielded approximately 300 ng of DNA. Such a total yield is currently insufficient for many high-throughput genome screening applications, necessitating whole genome amplification of all or most individuals in a population prior to genotyping.

Item Type:	Article
Additional Information:	Wilding, Craig S. Weetman, D. Steen, K. Donnelly, M. J.
Uncontrolled Keywords:	Anopheles gambiae; DNA quantification; mosquito; picogreen; spectrophotometry
Subjects:	QU Biochemistry > Cells and Genetics > QU 300 General works QX Parasitology > Insects. Other Parasites > QX 510 Mosquitoes QX Parasitology > Insects. Other Parasites > QX 650 Insect vectors QU Biochemistry > Genetics > QU 450 General Works QX Parasitology > Insects. Other Parasites > QX 515 Anopheles
Digital Object Identifer (DOI):	https://doi.org/10.1111/j.1744-7917.2009.01260.x
Depositing User:	Users 183 not found.
Date Deposited:	10 Mar 2010 14:17
Last Modified:	14 Dec 2021 10:13
URI:	https://archive.lstmed.ac.uk/id/eprint/411

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