Comparison of methods to extract PCR-amplifiable DNA from fruit, herbal and black teas

The success of polymerase chain reaction (PCR) assay depends on template deoxyribonucleic acid (DNA) being sufficient with respect to both quantity and quality. Some biological materials contain compounds which inhibit the functioning of DNA polymerase and thus need to be removed as part of the DNA extraction procedure. The aim of the present experiments was to optimise the process of DNA isolation from various types of black, fruit and herbal teas. A comparison was made between two cetyltrimethylammonium bromide (CTAB)-based protocols and two commercially available DNA purification kits. The yield and integrity of the extracted DNA were monitored both spectrophotometrically and using agarose gel electrophoresis. The presence/absence of inhibitors in the DNA preparations was checked by running quantitative real-time PCRs. The optimal protocol was deemed to be the CTAB method described in ISO 21571:2005, so this method is recommended for the routine sample analysis of tea products.

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Extraction and detection of animal deoxyribonucleic acid (DNA) species on lipsticks using Polymerase Chain Reaction (PCR) assay

International Journal of ChemTech Research ◽

10.20902/ijctr.2018.110632 ◽

2018 ◽

Keyword(s):

Polymerase Chain Reaction ◽

Deoxyribonucleic Acid ◽

Chain Reaction ◽

Pcr Assay ◽

Polymerase Chain

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Identifikasi Simplisia yang Dijual sebagai Strychnos ligustrina Bl. di Pasar Tradisional Surabaya dengan Metode Random Amplified Polymorphic DNA (RAPD)

Journal of Biological Researches ◽

10.23869/bphjbr.11.1.20054 ◽

2005 ◽

Vol 11 (1) ◽

pp. 19-24

Author(s):

Oeke Yunita ◽

Angelica Kresnamurti

Keyword(s):

Cetyltrimethylammonium Bromide ◽

Molecular Level ◽

Random Amplified Polymorphic Dna ◽

Local Market ◽

Chain Reaction ◽

Banding Patterns ◽

Traditional Market ◽

Ctab Method ◽

Polymerase Chain ◽

Rapd Method

Authentication of Strychnos ligustrina Bl. had been performed at molecular level (DNA) with Random Amplified Polymorphic DNA (RAPD) method, based on the amplification of random DNA fragments by Polymerase Chain Reaction (PCR) with a single arbitrary primer. The aim of this research was obtaining similar banding patterns between DNA of plant Strychnos ligustrina Bl. and DNA of its lignum on local market. Strychnos ligustrina Bl. was determined by UPT Balai Konservasi Tumbuhan Kebun Raya Purwodadi and plants sold as Strychnos ligustrina Bl. were collected as lignum from traditional market at Wonokromo, Rungkut, Genteng, Benowo dan Pabean. DNA from these plants were extracted by modified Cetyltrimethylammonium bromide (CTAB) method and amplified by RAPD method. Amplification had been performed by primer OPO-4 had shown banding patterns on the gel electrophoresis which banding patterns were shown by Strychnos ligustrina Bl. and plants sold as Strychnos ligustrina Bl. on Benowo. Based on this early result, we assume that plants sold as Strychnos ligustrina Bl. on Benowo has closely genetic relationship with Strychnos ligustrina Bl.

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Optimalization of DNA isolation from oral epithelial mucous cell with smear method

Padjadjaran Journal of Dentistry ◽

10.24198/pjd.vol21no3.14110 ◽

2009 ◽

Vol 21 (3) ◽

Author(s):

Saskia L. Nasroen ◽

Ani Melani Maskoen ◽

Agus Nurwiadh

Keyword(s):

Deoxyribonucleic Acid ◽

Dna Isolation ◽

Mucous Cell ◽

Genetic Material ◽

The Body ◽

Isolation Method ◽

Chain Reaction ◽

Polymerase Chain ◽

Living Organisms ◽

Oral Epithelial

Deoxyribonucleic acid (DNA) is a genetic material which is found in all living organisms. On the human cell or eukaryotes cell, the DNA is found in the nucleus cell and the mitochondria. The DNA arrangement on each cell in human body is the same, that is why, for the analysis meaning, DNA can be isolated from any cell in the body. The source of DNA to be analyzed usually coming from the blood sample by an injection method, such a way resulting in pain and bringing about constraint. Therefore, a study was carried out to look for an alternative of DNA isolation. The aim of this experimental study was to get an optimal DNA isolation method by using oral mucous smear method with a purpose to get a quick and easy DNA isolation. The investigation materials were in the form of samples which were taken from the oral epithelial mucous cells out of three different subjects. The epithelial cells were obtained by the oral mucous smear method which in a variation of two, four and six times of smear applications, respectively. The DNA was then isolated using buffer extraction method. The concentrations of DNA were measured by using ultraviolet spectrophotometer at 260 nm wavelength. The results of DNA isolation were analyzed by Polymerase Chain Reaction (PCR) technique. The optimal DNA isolation could be analyzed by PCR technique. The experimental results show that from three different subjects of study, DNA can be isolated optimally by oral mucous smear method with six times of smear applications.

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A fast CTAB method of human DNA isolation for polymerase chain reaction applications

Biochemical Education ◽

10.1016/s0307-4412(97)00122-2 ◽

1997 ◽

Vol 25 (4) ◽

pp. 233-235 ◽

Cited By ~ 8

Author(s):

John C. Thomas ◽

Rami Khoury ◽

Chris K. Neeley ◽

Ann M. Akroush ◽

Elizabeth C. Davies

Keyword(s):

Polymerase Chain Reaction ◽

Dna Isolation ◽

Chain Reaction ◽

Human Dna ◽

Ctab Method ◽

Polymerase Chain

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Polymerase chain reaction amplification and typing of rotavirus in environmental samples

Water Science & Technology ◽

10.2166/wst.1995.0643 ◽

1995 ◽

Vol 31 (5-6) ◽

pp. 371-374 ◽

Cited By ~ 5

Author(s):

R. Gajardo ◽

R. M. Pintó ◽

A. Bosch

Keyword(s):

Polymerase Chain Reaction ◽

Environmental Samples ◽

Polymerase Chain Reaction Amplification ◽

Rt Pcr ◽

Chain Reaction ◽

Pcr Assay ◽

Group A ◽

Reaction Amplification ◽

Stool Specimens ◽

Polymerase Chain

A reverse transcription polymerase chain reaction (RT-PCR) assay is described that has been developed for the detection and serotyping of group A rotavirus in stool specimens and concentrated and non-concentrated sewage specimens.

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Analysis and validation of a highly sensitive one-step nested quantitative real-time polymerase chain reaction assay for specific detection of severe acute respiratory syndrome coronavirus 2

Virology Journal ◽

10.1186/s12985-020-01467-y ◽

2020 ◽

Vol 17 (1) ◽

Author(s):

Yang Zhang ◽

Chunyang Dai ◽

Huiyan Wang ◽

Yong Gao ◽

Tuantuan Li ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Real Time ◽

Quantitative Polymerase Chain Reaction ◽

Clinical Samples ◽

Chain Reaction ◽

Pcr Assay ◽

Qrt Pcr ◽

Highly Sensitive ◽

One Step ◽

Polymerase Chain

Abstract Background Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30 to 60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. Method In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. Result The limit of detection (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI: 363.23–1145.69) for ORF1ab and 528.1 (95% CI: 347.7–1248.7) for N, 401.8 (95% CI: 284.8–938.3) for ORF1ab and 336.8 (95% CI: 244.6–792.5) for N, and 194.74 (95% CI: 139.7–430.9) for ORF1ab and 189.1 (95% CI: 130.9–433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. Conclusion In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.

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