Protocol for extraction of genomic DNA from swine solid tissues

Biase, Fernando H.; Franco, M. M.; Goulart, Luiz Ricardo; Antunes, Robson Carlos

doi:10.1590/s1415-47572002000300011

Cited by 61 publications

(28 citation statements)

References 2 publications

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“…Although milk is animal friendly and a routine source in the dairy industry (D'Angelo et al, 2007), inhibitors in milk such as fats and proteins render it difficult source for extracting high quantity and quality DNA (Lipkin et al, 1993;Amills et al, 1997;Murphy et al, 2002;Feligini et al, 2005;Cremonesi et al, 2006). To date, different commercial kits for improved DNA extraction are available for many kinds of tissues except for milk (Biase et al, 2002;Studer et al, 2008;O'Grady et al, 2008;Gao et al, 2011).…”

Section: Introductionmentioning

confidence: 99%

Comparison of methods for high quantity and quality genomic DNA extraction from raw cow milk

Usman¹,

Yu²,

Liu³

et al. 2014

Genet. Mol. Res.

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ABSTRACT. Isolation of sufficient quantities of high quality DNA is a prerequisite for molecular studies. Milk somatic cells can be used; however, inhibitors such as fats and proteins make milk a difficult medium for extracting large amounts of quality DNA. We optimized, evaluated and compared three methods, Modified Nucleospin Blood Kit method, Modified TianGen Kit method and Phenol-Chloroform method for genomic DNA extraction from bovine milk. Individual cows' milk and bulk milk samples were collected from a China agricultural university dairy farm. Genomic DNA extracted from each milk sample by the three methods was evaluated for quantity and purity by spectrophotometry and gel electrophoresis, as well as PCR and sequencing. All the three methods were found suitable for genomic DNA isolation from bovine milk, PCR applications, and sequencing. Comparing the three methods, we found that the Modified Nucleospin Blood Kit method was significantly better than the Phenol-Chloroform method in terms of quantity as well as quality (amount, concentration, 260/280nm and 260/230nm absorbance ratio), whereas, the Modified TianGen Kit method was more efficient than the Phenol-Chloroform method and cheaper than the Modified Nucleospine Blood Kit method; it yielded reasonably good quantities of good quality DNA and would be suitable for large-scale genotyping of lactating cows.

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Section: Introductionmentioning

confidence: 99%

Comparison of methods for high quantity and quality genomic DNA extraction from raw cow milk

Usman¹,

Yu²,

Liu³

et al. 2014

Genet. Mol. Res.

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“…The first challenge is the reduction of secondary chemical reactions (including oxidation) in the initial crude tissue extract, which otherwise could lead to loss of DNA yield; another is the lack of a universal extraction protocol suitable for different organisms because of the differences in compounds between them (Kotchomo and Gachomo, 2009). Previously, genomic DNA purification methods included phenol-chloroform-based approaches, with a popular method described by Sambrook et al (1989), using Chelex-100 (Walsh et al, 1991;Roberge et al, 1997;Yue and Orban, 2005) and methods based on grinding tissue in liquid nitrogen (Biase et al, 2002). These methods have been used effectively, but are time-consuming, accrue a storage and handling cost, and use hazardous reagents that require special handling and disposal mechanisms.…”

Section: Resultsmentioning

confidence: 99%

A simplified universal genomic DNA extraction protocol suitable for PCR

Wang¹,

Wang²,

Zhang³

et al. 2011

Genet. Mol. Res.

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ABSTRACT. Conventional genomic DNA extraction protocols need expensive and hazardous reagents for decontamination of phenolic compounds from the extracts and are only suited for certain types of tissue. We developed a simple, time-saving and cost-efficient method for genomic DNA extraction from various types of organisms, using relatively innocuous reagents. The protocol employs a single purification step to remove contaminating compounds, using a silica column and a non-hazardous buffer, and a chaotropic-detergent lysing solution that hydrolyzes RNA and allows the selective precipitation of DNA from cell lysates. We used this system to extract genomic DNA from different tissues of various organisms, including algae (Dunaliella salina), human peripheral blood, mouse liver, Escherichia coli, and Chinese hamster ovary cells. Mean DNA yields were 20-30 μg/cm 3 from fresh tissues (comparable to yields given by commercial extraction kits), and the 260/280 nm absorbance ratio was 1.8-2.0, demonstrating a good degree of purity. The extracted DNA was successfully used in PCR, restriction enzyme digestion and for recombinant selection studies.

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“…The suspension was incubated at 55°C for 3 hours. For precipitation of undissolved debris and proteins, 300 μl of 6 M NaCl was added to the suspension and kept at 4°C for 15 min, prior to centrifugation at 13,000 g for 15 min, after which the supernatants were transferred to new Eppendorf tubes (Biase et al, 2002). Then, the samples were extracted with phenolchloroform-isoamyl alcohol (25:24:1).…”

Section: Methodsmentioning

confidence: 99%

Molecular detection of Neospora caninum in house sparrows (Passer domesticus) in Iran

et al. 2015

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Neospora caninum is an intracellular protozoan parasite with a wide range of intermediate bird hosts. There is little information describing the prevalence and genetic characterization of N. caninum in bird hosts worldwide and in Iran. In this study, a total of 217 brain samples of house sparrow (Passer domesticus) were examined for N. caninum presence by nested polymerase chain reaction targeting the Nc-5 gene. N. caninum DNA was detected in 3.68% (8/217) of sparrows. Sequencing of the Nc5 genomic DNA revealed 97-99% of similarity with N. caninum sequences deposited in Genbank. To our knowledge, this study is the first molecular evidence of N. caninum DNA in bird hosts in Iran. The results of this study highlight the role of the house sparrow (Passer domesticus) in maintaining and spreading N. caninum infection to canines in the feral and domestic environment.

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Protocol for extraction of genomic DNA from swine solid tissues

Cited by 61 publications

References 2 publications

Comparison of methods for high quantity and quality genomic DNA extraction from raw cow milk

Comparison of methods for high quantity and quality genomic DNA extraction from raw cow milk

A simplified universal genomic DNA extraction protocol suitable for PCR

Molecular detection of Neospora caninum in house sparrows (Passer domesticus) in Iran

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