A simple plant high-molecular-weight DNA extraction method suitable for single-molecule technologies

Li, Zhigang; Parris, Stephen; Saski, Christopher

doi:10.1186/s13007-020-00579-4

Cited by 34 publications

(30 citation statements)

References 3 publications

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“…Although abaca is a close relative of banana, their leaves' brous nature and higher phenolic content has rendered DNA extraction more di cult. Several methods to isolate DNA from various plant tissues are available, but these methods are time consuming and produce inconsistent yield and quality [20,21]. Since the rst application of CTAB-based extraction method for plants [22], there have been variations depending on the plant species.…”

Section: Dna Extraction Methods Comparisonmentioning

confidence: 99%

“…Leaf tissue samples were freshly cut from the second to the youngest leaf stalk of the healthy abaca plant prior to start of the DNA extraction process thus phenolic content may be relatively low and oxidation of phenols may not have started yet upon cutting. This highlights the importance of selecting fresh leaf samples from young healthy plants and its immediate use in DNA extraction experiments to prevent oxidation of phenols [14,21]. Moreover, the addition of liquid nitrogen allowed rapid disruption of a tissue into powder form at low temperature [29], thus oxidation of available plant phenolic content could have been minimized due to low chemical and biological activity at this temperature.…”

Section: Dna Extraction Methods Comparisonmentioning

confidence: 99%

“…Moreover, the addition of liquid nitrogen allowed rapid disruption of a tissue into powder form at low temperature [29], thus oxidation of available plant phenolic content could have been minimized due to low chemical and biological activity at this temperature. It is a common practice in plant DNA extraction that cell wall disruption to release nuclei and organelles are done in the presence of liquid nitrogen to prevent increased exposure time to air and warm temperature before the ground tissue is completely mixed with the extraction buffer [21].…”

Section: Dna Extraction Methods Comparisonmentioning

confidence: 99%

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Extraction of high molecular weight abaca DNA suitable for next-generation sequencing

Koh

Barbosa²,

Aquino

et al. 2020

Preprint

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Background The abaca (Musa textilis Née) is a fiber crop native to the Philippines with high economic value because of its fiber - the Manila hemp, known to be the strongest of all the natural fibers. DNA extraction in abaca is difficult due to its fibrous nature, high cellulose content and polyphenol compounds. Thus an optimized DNA extraction method is required for extracting high quality abaca DNA for next-generation sequencing applications. Results In this study, we have compared five different methods for the extraction of high molecular weight DNA from abaca leaves. The methods are the traditional CTAB method (Protocol 1), the CTAB with PVP method (Protocol 2), the CTAB with 0.3% β-mercaptoethanol method (Protocol 3), SDS-method (Protocol 4) and CTAB with Triton X-100 and PVP method (Protocol 5). Out of the five methods tested, traditional CTAB-method (Protocol 1), CTAB with 0.3% β-mercaptoethanol method (Protocol 3) and SDS-method (Protocol 4) have shown to be the most consistent in giving high molecular weight DNA with good yield and purity based on A260/A280 and A260/A230 absorption values. TissueLyserII was also utilized for homogenization for the three extraction protocols for applications in high-throughput DNA extraction. DNA from two abaca varieties were extracted using the CTAB with 0.3% β-mercaptoethanol method (Protocol 3) and were sent for NGS based on Illumina HiSeq platform having both passed the quality control for library preparation. Conclusion The CTAB with 0.3% β-mercaptoethanol method (Protocol 3) was found to be the simplest and most consistent method for extracting average yield DNA with high quality for NGS applications. The SDS-method (Protocol 4) was determined to have the shortest processing time and together with TissueLyserII is the most appropriate method for high-throughput extraction of abaca samples which will be useful for genotyping-by-sequencing (GBS) studies.

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Section: Dna Extraction Methods Comparisonmentioning

confidence: 99%

Section: Dna Extraction Methods Comparisonmentioning

confidence: 99%

Section: Dna Extraction Methods Comparisonmentioning

confidence: 99%

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Extraction of high molecular weight abaca DNA suitable for next-generation sequencing

Koh

Barbosa²,

Aquino

et al. 2020

Preprint

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“…The plant material was obtained from foundation stock of the cultivar 'Royal Royce' maintained by the UC Davis Strawberry Breeding Program. DNA was isolated as previously described 30 . The genomic DNA was larger than required for HiFi library production and was sheared using the Megaruptor 3 targeting a size distribution centered around 22 kb.…”

Section: F × Ananassa 'Royal Royce' Sample Acquisition Dna Extractimentioning

confidence: 99%

Highly accurate long-read HiFi sequencing data for five complex genomes

Hon

Mars

Young

et al. 2020

Preprint

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The PacBio ® HiFi sequencing method yields highly accurate long-read sequencing datasets with read lengths averaging 10-25 kb and accuracies greater than 99.5%. These accurate long reads can be used to improve results for complex applications such as single nucleotide and structural variant detection, genome assembly, assembly of difficult polyploid or highly repetitive genomes, and assembly of metagenomes. Currently, there is a need for sample data sets to both evaluate the benefits of these long accurate reads as well as for development of bioinformatic tools including genome assemblers, variant callers, and haplotyping algorithms. We present deep coverage HiFi datasets for five complex samples including the two inbred model genomes Mus musculus and Zea mays, as well as two complex genomes, octoploid Fragaria × ananassa and the diploid anuran Rana muscosa. Additionally, we release sequence data from a mock metagenome community. The datasets reported here can be used without restriction to develop new algorithms and explore complex genome structure and evolution. Data were generated on the PacBio Sequel II System.

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“…All these compounds affect the quality and quantity of isolated DNA and reduce the ampli cation of samples in polymerase chain reaction (PCR) [6]. For high-quality results in high throughput sequencing (HTS) techniques and other molecular techniques, extraction of high quality and pure DNA is essential [7,8].…”

Section: Introductionmentioning

confidence: 99%

Comparison of genomic DNA extraction methods to obtain high DNA quality from barnyard grass (Echinochloa colona)

Salgotra

Chauhan

2020

Preprint

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Background: The study of weed genomics is important for the effective management of weeds to enhance crop yield. A rapid, inexpensive and high quality DNA extraction is needed for genomic and other molecular studies. Here, we describe the protocols for DNA extraction from two different parts of the Echinochloa colona plant using modified cetyltrimethylammonium bromide (CTAB) and a commercial kit.Results: In the study, it was observed that the DNA extracted from plant leaf tissues and dry seeds with a modified CTAB protocol was of good quality, with no contaminations of polysaccharides and essential oils. Quality of DNA extracted from dry seeds was comparable with that of plant leaves under both protocols. The extracted DNA from both plant parts was successfully amplified by PCR using the EPSPS microsatellite marker. Compared to the protocol of DNA extracted from leaf tissue, dry seeds will save time and other valuable resources. Moreover, the same protocols can be implemented for the extraction of high-quality DNA for molecular studies in other plant species where a large amount of polysaccharides, secondary metabolites and essential oils are present.Conclusions: Modified methods of DNA extraction from dry seeds are efficient and time-saving which can be used in genotypic and other molecular approaches. High-quality DNA can be isolated from plant leaf tissues using modified CTAB and commercial kits, however, DNA extracted from dry seeds will save time and other valuable resources.

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A simple plant high-molecular-weight DNA extraction method suitable for single-molecule technologies

Cited by 34 publications

References 3 publications

Extraction of high molecular weight abaca DNA suitable for next-generation sequencing

Extraction of high molecular weight abaca DNA suitable for next-generation sequencing

Highly accurate long-read HiFi sequencing data for five complex genomes

Comparison of genomic DNA extraction methods to obtain high DNA quality from barnyard grass (Echinochloa colona)

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