SABIIA - Sistema Aberto e Integrado de Informação em Agricultura (Sabiia)

Silva,Gildo Almeida da; Bernardi,Taís Letícia; Schaker,Patrícia Dayane Carvalho; Menegotto,Morgana; Valente,Patricia.

The purpose of this work was to study a rapid yeast DNA extraction by boiling and freeze-thawing processes without using chemical reagents or any purification procedures, to obtain a high grade PCR-product. A specific DNA fragment of the 18S region of Dekkera bruxellensis and Saccharomyces cerevisiae was chosen. The described boiling and freeze-thawing protocols generated the PCR-grade product preparations and could be used to process many samples. The amplification of the fragments could be observed after 30 and 35 cycles. These processes of extraction without using any kind of chemical reagents, especial water, and purification procedures proved to be efficient, reproducible, simple, fast, and inexpensive.

Direct RAPD evaluation of bacteria without conventional DNA extraction

Palavras-chave: Boiling; Freeze-thawing; DNA extraction; Dekkera bruxellensis; Saccharomyces cerevisiae.

Ano: 2012

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132012000200020

Araújo,Welington Luiz; Angellis,Derlene Attili de; Azevedo,João Lúcio.

The present work reports successful DNA amplification of Pantoea agglomerans and Bacillus pumilus through Random Amplified Polymorphic DNA (RAPD). For this, template DNA was obtained without conventional DNA extraction. The procedure was as follows: cultures grown for 20 hours in 5 mL LB medium were centrifuged and the resulting preparation was suspended in TE buffer. After boiling, the cell suspension was diluted and 2.0 µl were used in reactions of 15 µl. The results showed no significant differences among the RAPD profile of the PCR reactions derived from the boiling and phenol extraction methods, suggesting the utilization of this method for genetic population analysis.

An efficient and rapid DNA minipreparation procedure suitable for PCR/SSR and RAPD analyses in tropical forest tree species

Palavras-chave: Bacteria; Boiling method; DNA extraction; RAPD.

Ano: 2004

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132004000300006

Alzate-Marin,Ana Lilia; Guidugli,Marcela Corbo; Soriani,Hilda Hildebrand; Martinez,Carlos Alberto; Mestriner,Moacyr Antônio.

An efficient and rapid DNA minipreparation modified method for frozen samples was developed for five tropical tree species: Copaifera langsdorffii, Hymenaea courbaril, Eugenia uniflora, Tabebuia roseo alba and Cariniana estrellensis. This procedure that dispenses the use of liquid nitrogen, phenol and the addition of proteinase K, is an adaptation of the CTAB-based DNA extraction method. The modifications included the use of PVP to eliminate the polyphenols, only one chloroform-isoamyl alcohol step and the addition of RNase immediately after extraction with chloroform. The yields of the DNA samples ranged from 25.7 to 42.1 µg from 100 mg leaf tissue. The DNA samples extracted by this method were successfully used for PCR (SSR and RAPD) analyses in these...

An Alternative and Rapid Method for the Extraction of Nucleic Acids from Ixodid Ticks by Potassium Acetate Procedure

Palavras-chave: Copaifera langsdorffii; Hymenaea courbaril; Eugenia uniflora; Tabebuia roseo alba; Cariniana estrellensis; DNA extraction.

Ano: 2009

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132009000500020

Rodríguez,Islay; Fraga,Jorge; Noda,Angel Alberto; Mayet,Milagro; Duarte,Yanisia; Echevarria,Eduardo; Fernández,Carmen.

Four variants of the potassium acetate procedure for DNA extraction from ixodid ticks at different stage of their life cycles were evaluated and compared with phenol-chloroform and ammonium hydroxide methods. The most rapid and most efficient variant was validated in the DNA extraction procedure from the engorged ticks collected from bovine, canine as well as from house ticks for the screening of Borrelia burgdorferi sensu lato, Anaplasma spp. and Babesia spp. The ammonium hydroxide procedure was used for non-engorged ticks. All the variants were efficient and allowed obtaining PCR-quality material according to the specific amplification of 16S rRNA gene fragment of the original tick. DNA extracted from the ticks under the study was tested by multiplex PCR...

Development of an Extraction Protocol for Fetal Sexing from the Fourth Week of Gestation by Real Time PCR

Palavras-chave: Ticks; PCR; DNA extraction.

Ano: 2014

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132014000400542

Oliveira,Edwin José Torres de; Rabacow,Ana Paula Maluf; Figueiredo,Bruno Pereira de; Ishikawa,Raíssa Borges; Pessatto,Lucas Roberto; Pelizaro,Bruno Ivo; Figueiredo,Helder Pereira de; Oliveira,Rodrigo Juliano.

Abstract Low number of fetal cells in maternal blood limited the use of fetal materials in diagnostic and clinical applications. This research developed a technology which allowed the extraction of fetal DNA by a non-invasive method that offers no risk to the mother or fetus. A total of 132 pregnant women participated in this inquiry. The DNA extraction was performed employing an in-house method based on guanidine thiocyanate and magnetic bead. For the amplification it was used the Quantifier Y Kit™. The fetal sexing analysis of the 132 pregnant women were 100% in agreement with the ultrasound. Sensitivity and specificity for detection of Y chromosome sequences was possible using Real-Time Polymerase Chain Reaction since the 4th week of gestation. This...

Urine as a promising sample for Leishmania DNA extraction in the diagnosis of visceral leishmaniasis - a review

Palavras-chave: Fetal-sexing; DNA extraction; RtPCR..

Ano: 2019

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132019000100416

BJID

Bezerra,Gilberto Silva Nunes; Barbosa Júnior,Walter Lins; Silva,Elis Dionísio da; Leal,Nilma Cintra; Medeiros,Zulma Maria de.

ABSTRACT Visceral leishmaniasis is a serious and debilitating infection with high fatality rate in tropical and subtropical countries. As clinical symptoms of visceral leishmaniasis are not so specific, confirmatory diagnostic methods with high sensitivity and specificity are needed. Noninvasive methods have been developed using urine as a clinical sample for visceral leishmaniasis diagnosis. In fact, there is a clear correlation between kidney impairment and Leishmania DNA in urine. However, it has been proved that Leishmania nucleic acid may also be isolated from patients without any sign of renal involvement. Even though urine has become a promissing biological sample, it is still not widely used due to several issues, such as (i) incomprehension of the...

Use of a cysteine proteinase from Carica candamarcensis as a protective agent during DNA extraction

Palavras-chave: Visceral leishmaniasis; Renal involvement; Diagnosis; Urine sample; DNA extraction.

Ano: 2019

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1413-86702019000200111

BJMBR

Genelhu,M.S.; Zanini,M.S.; Veloso,I.F.; Carneiro,A.M.D.; Lopes,M.T.P.; Salas,C.E..

We describe the use of a plant cysteine proteinase isolated from latex of Carica candamarcensis as a protective agent during isolation of bacterial DNA following growth in culture of these cells. Between 100 to 720 units of proteinase (1 µg = 6 units) afforded good DNA protection when incubated with various kinds of microorganisms. Agarose gel electrophoresis showed that the resulting DNA was similar in size to DNA preparations obtained by treatment with proteinase K. The viability of the resulting material was checked by PCR amplification using species-specific primers. After standing at room temperature (25oC) for 35 days, the enzyme lost 10% of its initial activity. The enzyme stability and good yield of DNA suggest the use of this proteinase as an...

Tipo: Info:eu-repo/semantics/other

Palavras-chave: Cysteine proteinase; Proteinase K; C. candamarcensis; DNA extraction.

Ano: 1998

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X1998000900005

Enhancement of soil DNA extraction by the use of a hand held mixer

Costa,Jefferson Luis da Silva; Oliveira,Virgínia Carla de.

The efficiency of soil DNA extraction using a bead beater homogenizer (Biospec Products - Germany) was compared to that obtained with a hand held mixer (Moulinex - Brazil). The hand held mixer costs 100 times less and extracted seven times more crude DNA than the bead beater.

Tipo: Info:eu-repo/semantics/other

Palavras-chave: Molecular tools; DNA extraction.

Ano: 2003

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822003000400004

Comparison of different protocols for the extraction of microbial DNA from reef corals

Santos,H.F.; Carmo,F.L.; Leite,D.C.A.; Jesus,H.E.; De Carvalho Maalouf,P.; Almeida,C.; Soriano,A.U.; Altomari,D.; Suhett,L.; Vólaro,V.; Valoni,E.; Francisco,M.; Vieira,J.; Rocha,R.; Sardinha,B.L.; Mendes,L.B.; João,R.R.; Lacava,B.; Jesus,R.F.; Sebastian,G.V.; Pessoa,A.; van Elsas,J.D.; Rezende,R.P.; Pires,D.O.; Duarte,G.; Castro,C.B.; Rosado,A.S.; Peixoto,R.S..

This study aimed to test different protocols for the extraction of microbial DNA from the coral Mussismilia harttii. Four different commercial kits were tested, three of them based on methods for DNA extraction from soil (FastDNA SPIN Kit for soil, MP Bio, PowerSoil DNA Isolation Kit, MoBio, and ZR Soil Microbe DNA Kit, Zymo Research) and one kit for DNA extraction from plants (UltraClean Plant DNA Isolation Kit, MoBio). Five polyps of the same colony of M. harttii were macerated and aliquots were submitted to DNA extraction by the different kits. After extraction, the DNA was quantified and PCR-DGGE was used to study the molecular fingerprint of Bacteria and Eukarya. Among the four kits tested, the ZR Soil Microbe DNA Kit was the most efficient with...

Evaluation of DNA extraction protocols for Brucella abortus pcr detection in aborted fetuses or calves born from cows experimentally infected with strain 2308

Palavras-chave: Coral; Microbial diversity; DNA extraction.

Ano: 2012

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822012000200012

Matrone,M.; Keid,L.B.; Rocha,V.C.M.; Vejarano,M.P.; Ikuta,C.Y.; Rodriguez,C.A.R.; Ferreira,F.; Dias,R.A.; Ferreira Neto,J.S.

The objective of the present study was to improve the detection of B. abortus by PCR in organs of aborted fetuses from infected cows, an important mechanism to find infected herds on the eradication phase of the program. So, different DNA extraction protocols were compared, focusing the PCR detection of B. abortus in clinical samples collected from aborted fetuses or calves born from cows challenged with the 2308 B. abortus strain. Therefore, two gold standard groups were built based on classical bacteriology, formed from: 32 lungs (17 positives), 26 spleens (11 positives), 23 livers (8 positives) and 22 bronchial lymph nodes (7 positives). All samples were submitted to three DNA extraction protocols, followed by the same amplification process with the...

Assessment of the quality of dna extracted by two techniques from Mycobacterium tuberculosis for fast molecular identification and genotyping

Palavras-chave: Bovine; Brucellosis; Abortion; PCR; DNA extraction.

Ano: 2009

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822009000300010

Miyata,Marcelo; Santos,Adolfo Carlos Barreto; Mendes,Natália Helena; Cunha,Eunice Atsuko; Melo,Fernando Augusto Fiúza de; Leite,Clarice Queico Fujimura.

We report a comparative study of two DNA extraction techniques, thermolysis and chemical lysis (CTAB), for molecular identification and genotyping of M. tuberculosis. Forty DNA samples were subjected to PCR and the results demonstrated that with thermolysis it is possible to obtain useful data that enables fast identification and genotyping.

Comparison of DNA extraction protocols for microbial communities from soil treated with biochar

Palavras-chave: DNA extraction; Mycobacterium tuberculosis; Thermolysis; CTAB.

Ano: 2011

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822011000200045

Simple and robust DNA extraction method for the large-scale analysis of genotypes containing high polyphenolic content, such as landracesof Solanum tuberosum and Zea mays

Leite,D.C.A.; Balieiro,F.C.; Pires,C.A.; Madari,B.E.; Rosado,A.S.; Coutinho,H.L.C.; Peixoto,R.S..

Many studies have evaluated the effects of biochar application on soil structure and plant growth. However, there are very few studies describing the effect of biochar on native soil microbial communities. Microbial analysis of environmental samples requires accurate and reproducible methods for the extraction of DNA from samples. Because of the variety among microbial species and the strong adsorption of the phosphate backbone of the DNA molecule to biochar, extracting and purifying high quality microbial DNA from biochar-amended soil is not a trivial process and can be considerably more difficult than the extraction of DNA from other environmental samples. The aim of this study was to compare the relative efficacies of three commercial DNA extraction...

Tipo: Info:eu-repo/semantics/other

Palavras-chave: Biochar; DNA extraction; PCR-DGGE; Microbial communities; DNA purity indices.

Ano: 2014

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822014000100023

Ciencia e Investigación Agraria

Barra,Macarena; Salazar,Erika; Beltrán,María; Sagredo,Boris.

The extraction of high-quality DNA is essential for studies conducted at the molecular level in species with an abundance of contaminants in their tissues, such as some landraces of potato and maize, in which it is difficult to extract good-quality genomic DNA. Compounds such as polyphenols interfere with the amplification of DNA during polymerase chain reaction (PCR), which makes it difficult to conduct PCR-based studies ofmolecular markers. This article describes a simple and robust protocol for DNA isolation that was applied to potato and maize landraces and resulted in a high DNA concentration with excellent purity and quality. This method is based on the method of Krizman et al. (2006) with some modifications and was tested on potato leaves and young...

Tipo: Journal article

Palavras-chave: Activated charcoal; DNA extraction; Genomic DNA; Polyphenols.

Ano: 2012

URL: http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0718-16202012000300019

Comparison of nine DNA extraction methods for the diagnosis of bovine tuberculosis by real time PCR

Ciência Rural

Moura,André; Hodon,Mikael Arrais; Soares Filho,Paulo Martins; Issa,Marina de Azevedo; Oliveira,Ana Paula Ferreira de; Fonseca Júnior,Antônio Augusto.

ABSTRACT: Bovine tuberculosis is an infectious disease with a high impact on the cattle industry, particularly in developing countries. PCR is a very sensitive method for detection of infectious agents, but the sensitivity of molecular diagnosis is largely dependent on the efficiency of the DNA extraction methods. The objective of this study was to evaluate DNA extraction methods for direct detection of Mycobacterium bovis in bovine tissue. Nine commercial kits for DNA extraction were evaluated when combined with two real time PCRs. The DNeasy Blood & Tissue Kit from QIAGEN showed better performance and sensitivity followed by the DNA Mini Kit RBC and FTA Elute Micro Card. Results suggested that, even when the analytical sensitivity of the qPCR is...