Benchmarking ultra-high molecular weight DNA preservation methods for long-read and long-range sequencing.
Gigascience
; 112022 08 10.
Article
em En
| MEDLINE
| ID: mdl-35946988
ABSTRACT
BACKGROUND:
Studies in vertebrate genomics require sampling from a broad range of tissue types, taxa, and localities. Recent advancements in long-read and long-range genome sequencing have made it possible to produce high-quality chromosome-level genome assemblies for almost any organism. However, adequate tissue preservation for the requisite ultra-high molecular weight DNA (uHMW DNA) remains a major challenge. Here we present a comparative study of preservation methods for field and laboratory tissue sampling, across vertebrate classes and different tissue types.RESULTS:
We find that storage temperature was the strongest predictor of uHMW fragment lengths. While immediate flash-freezing remains the sample preservation gold standard, samples preserved in 95% EtOH or 20-25% DMSO-EDTA showed little fragment length degradation when stored at 4°C for 6 hours. Samples in 95% EtOH or 20-25% DMSO-EDTA kept at 4°C for 1 week after dissection still yielded adequate amounts of uHMW DNA for most applications. Tissue type was a significant predictor of total DNA yield but not fragment length. Preservation solution had a smaller but significant influence on both fragment length and DNA yield.CONCLUSION:
We provide sample preservation guidelines that ensure sufficient DNA integrity and amount required for use with long-read and long-range sequencing technologies across vertebrates. Our best practices generated the uHMW DNA needed for the high-quality reference genomes for phase 1 of the Vertebrate Genomes Project, whose ultimate mission is to generate chromosome-level reference genome assemblies of all â¼70,000 extant vertebrate species.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Dimetil Sulfóxido
/
Benchmarking
Tipo de estudo:
Guideline
Limite:
Animals
Idioma:
En
Revista:
Gigascience
Ano de publicação:
2022
Tipo de documento:
Article
País de afiliação:
Canadá