DNA Isolation and Amplification from Formaldehyde-Fixed Animal Tissues Rich in Mucopolysaccharides, Pigments, and Chitin

Palmer, Aparna D.-N.

doi:10.1080/10826060802589635

Cited by 5 publications

(3 citation statements)

References 10 publications

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“…Of the few genetic studies of formalin-fixed museum specimens, most have targeted nuclear (Bibi et al, 2015;Joshi et al,2013;Lutterschmidt et al, 2010;Palmer, 2009;Scatena & Morielle-Versute, 2008;Shiozaki et al, 2021) and high copy mitochondrial (Appleyard et al, 2021;Boyle et al, 2004;Shedlock et al, 1997) (Hykin et al,2015) and king cobra (Straube et al, 2021) has yielded sufficient coverage to reconstruct entire mitochondrial genomes. Using hot alkaline extraction, whole genomes were recovered for the bioluminescent bacterial symbionts contained within light organs of formalin-preserved cardinalfish (Gould et al,2020).…”

Section: Introductionmentioning

confidence: 99%

“…Of the few genetic studies of formalin‐fixed museum specimens, most have targeted nuclear (Bibi et al, 2015; Joshi et al,2013; Lutterschmidt et al, 2010; Palmer, 2009; Scatena & Morielle‐Versute, 2008; Shiozaki et al, 2021) and high copy mitochondrial (Appleyard et al, 2021; Boyle et al, 2004; Shedlock et al, 1997) loci via PCR amplification due to the difficulty and unpredictability of nuclear DNA extraction. There are few examples of broader‐scale genomic sequencing of formalin‐fixed museum specimens and none have recovered whole vertebrate genomes.…”

Section: Introductionmentioning

confidence: 99%

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Unlocking inaccessible historical genomes preserved in formalin

Hahn¹,

Alexander²,

Grealy³

et al. 2021

Molecular Ecology Resources

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Museum specimens represent an unparalleled record of historical genomic data.However, the widespread practice of formalin preservation has thus far impeded genomic analysis of a large proportion of specimens. Limited DNA sequencing from formalin-preserved specimens has yielded low genomic coverage with unpredictable success. We set out to refine sample processing methods and to identify specimen characteristics predictive of sequencing success. With a set of taxonomically diverse specimens collected between 1962 and 2006 and ranging in preservation quality, we compared the efficacy of several end-to-end whole genome sequencing workflows alongside a k-mer-based trimming-free read alignment approach to maximize mapping of endogenous sequence. We recovered complete mitochondrial genomes and up to 3× nuclear genome coverage from formalin-preserved tissues. Hot alkaline lysis coupled with phenol-chloroform extraction out-performed proteinase K digestion in recovering DNA, while library preparation method had little impact on sequencing success. The strongest predictor of DNA yield was overall specimen condition, which additively interacts with preservation conditions to accelerate DNA degradation. Here, we demonstrate a significant advance in capability beyond limited recovery of a small number of loci via PCR or target-capture sequencing. To facilitate strategic selection of suitable specimens for genomic sequencing, we present a decision-making framework that utilizes independent and nondestructive assessment criteria. Sequencing of formalin-preserved specimens will contribute to a greater understanding of temporal trends in genetic adaptation, including those associated with a changing climate. Our work enhances the value of museum collections worldwide by unlocking genomes of specimens that have been disregarded as a valid molecular resource.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Unlocking inaccessible historical genomes preserved in formalin

Hahn¹,

Alexander²,

Grealy³

et al. 2021

Molecular Ecology Resources

View full text Add to dashboard Cite

show abstract

“…Of the few genetic studies of formalin-fixed museum specimens, most have targeted nuclear (33)(34)(35)(36)(37) and high copy mitochondrial (20,38,39) loci via PCR amplification due to the difficulty and unpredictability of nuclear DNA extraction. There are few examples of broaderscale genomic sequencing of formalin-fixed museum specimens and none have recovered whole vertebrate genomes.…”

Section: Introductionmentioning

confidence: 99%

Unlocking inaccessible historical genomes preserved in formalin

Hahn

Alexander

Grealy

et al. 2021

Preprint

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Background: Museum specimens represent an unparalleled record of historical genomic data. However, the wide-spread practice of formalin preservation has thus far impeded genomic analysis of a large proportion of specimens. Limited DNA sequencing from formalin-preserved specimens has yielded low genomic coverage with unpredictable success. We set out to refine sample processing methods and to identify specimen characteristics predictive of sequencing success. With a set of taxonomically diverse specimens collected between 1936 and 2015 and ranging in preservation quality, we compared the efficacy of several end-to-end whole genome sequencing workflows alongside a k-mer-based trimming-free read alignment approach to maximize mapping of endogenous sequence. Results: We recovered complete mitochondrial genomes and up to 3X nuclear genome coverage from formalin-fixed tissues. Hot alkaline lysis coupled with phenol-chloroform extraction out-performed proteinase K digestion in recovering DNA, while library preparation method had little impact on sequencing success. The strongest predictor of DNA yield was overall specimen condition, which additively interacts with preservation conditions to accelerate DNA degradation. Conclusions: We demonstrate a significant advance in capability beyond limited recovery of a small number of loci via PCR or target-capture sequencing. To facilitate strategic selection of suitable specimens for genomic sequencing, we present a decision-making framework that utilizes independent and non-destructive assessment criteria. Sequencing of formalin-fixed specimens will contribute to a greater understanding of temporal trends in genetic adaptation, including those associated with a changing climate. Our work enhances the value of museum collections worldwide by unlocking genomes of specimens that have been disregarded as a valid molecular resource.

show abstract