Optimizing the pure genomic DNA isolation procedure for Plectranthus amboinicus DNA – A prerequisite for further genomic Studies

The DNA isolation procedure for different plant groups have been studied and standardized. The isolation of pure genomic DNA is the most essential component for any type of molecular studies. The present work is aimed to identify suitable DNA markers for the amplification of P.amboinicus DNA becomes a great hurdle for DNA barcoding studies carried out by rbcL and matK primers Used in the members of Lamiaceae. To solve this problem, The DNA was extracted by three methods from fresh young leaf tissue of P.amboinicus. After the evaluationthe outcome of these methods, one most suitable modified method was selected for isolating DNA from young leaves of P.amboinicus and selected for suitable DNA barcoding markers for PCR amplification. The quality and quantity of DNAs are a prerequisite for genetic studies for a variety of plants including P.amboinicus. The quantity and quality of the DNA extracted by this method wasused for suitable DNA barcoding markers selection.

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Rapid and High Quality DNA Isolation from Origanum onites for RAPD and ISSR Analysis

Zeitschrift für Naturforschung C ◽

10.1515/znc-2008-7-821 ◽

2008 ◽

Vol 63 (7-8) ◽

pp. 595-598 ◽

Cited By ~ 2

Author(s):

Emel Sözen ◽

Ismail Poyraz

Keyword(s):

Genomic Dna ◽

Dna Isolation ◽

Leaf Tissue ◽

Isoamyl Alcohol ◽

Molecular Study ◽

Inter Simple Sequence Repeat ◽

Limiting Factor ◽

Hexadecyltrimethylammonium Bromide ◽

Genomic Dna Isolation ◽

Origanum Onites

Origanum onites is an economically important medicinal plant with high essential oil content. Lack of an appropriate DNA isolation procedure is a limiting factor for any molecular study of this plant. We have used a protocol for genomic DNA isolation based on a hexadecyltrimethylammonium bromide (CTAB) method described for other plant species. The method involves mortar grinding of leaf tissue, modified CTAB extraction using high salt concentrations and polyvinyl pyrrolidone, and successive isoamyl alcohol/chloroform extractions. The yield was approx. 20 μg DNA per 200 mg of initial fresh plant material. The genomic DNA obtained by this method was suitable to be used in restriction digests, inter simple sequence repeat (ISSR) and randomly amplified polymorphic DNA (RAPD) reactions. This extraction method should facilitate the molecular analysis of Origanum chemotypes.

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Extraction of high-quality genomic DNA and identification of different DNA barcoding markers for chickpea (Cicer arietinum L.)

Buletin Agroteknologi ◽

10.32663/ba.v1i1.1194 ◽

2020 ◽

Vol 1 (1) ◽

pp. 7

Author(s):

Umavathi Saraswathi ◽

Lakshmanan Mullainathan

Keyword(s):

Dna Barcoding ◽

Dna Extraction ◽

Genomic Dna ◽

Dna Isolation ◽

Genomic Library ◽

Pcr Amplification ◽

Slight Modification ◽

High Quality ◽

Universal Primer ◽

Genetic Studies

The genetic studies of individual plants, especially self-pollinated species like chickpea need to be evaluated at the DNA level with the help of molecular markers for identifying genetic variations among the plants. High-quality DNA extraction is a prerequisite for genetic studies. Extraction of intact genomic DNA with high – molecular mass is essential for the study of many molecular biology applications like Polymerase Chain Reaction, endonuclease restriction digestion, southern blot analysis, and also for the construction of a genomic library. Several plant DNA extraction methods are available, even though the DNA isolation methods that give good yield employing both quantity and quality is quite difficult especially for self-pollinated crops like a chickpea. This work was focused on developing a standard protocol for the extraction of genomic DNA and identifying different barcoding markers. The result revealed that the CTAB extraction method with slight modification in protocol had been optimized for DNA isolation. The purified DNA, which was isolated through the CTAB method, had excellent spectral qualities and is efficiently digested by a restriction endonuclease, and is found to be more suitable for long-fragment PCR amplification. DNA barcoding is considered as a promising tool because it provides a practical and standard identification of plants. The isolated DNA sample was processed with a classical DNA barcoding approach by amplifying and sequencing with a universal primer. According to the result, among the different barcoding markers studied, the RbcL and Mat K were found to given the best result for molecular species identification in chickpea.

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Uji Efektivitas Metode Isolasi DNA Genom Kopi Arabika (Coffea arabica L.) Asal Kabupaten Jayawijaya

JURNAL BIOLOGI PAPUA ◽

10.31957/jbp.10 ◽

2018 ◽

Vol 8 (1) ◽

Author(s):

Arsyam Mawardi ◽

Maria L. Simonapendi

Keyword(s):

Genomic Dna ◽

Dna Isolation ◽

Dna Amplification ◽

Initial Information ◽

Arabica Coffee ◽

Modified Method ◽

Ctab Method ◽

Young Leaves ◽

Amplification Technique ◽

The Difference

Genetic substance of DNA has many functions as a basic component of the organism. DNA can be obtained directly through the isolation of DNA. Isolation of genomic DNA Wamena arabica coffee is done by treating the young leaves to get DNA extract. This reasearch is intended to provide scientific contributions in an effort to screen the best methods of DNA isolation, including a modified extraction of Zheng et al method (2005), a modified method of Doyle and Doyle (1990), and method of George and Khan modificaation (2008) or CTAB method. All of methods were tested on Arabica coffee Jayawijaya Wamena. Testing was done by looking at the difference in quality and quantity of products in the form of genomic DNA concentration and DNA thickness, comparing with the marker DNA then electrophoresis on agarose gel. From the results of testing the effectiveness of three types of isolation methods, it was found that the method of George and Khan (2008) or CTAB genomic DNA produce the best quality than other methods. In terms of quantity, the criteria in the form of DNA concentrations ranging from 100 ng λ-DNA/ml, λ-DNA 50 ng/ml, λ-DNA 10 ng/ml. Concentration of genomic DNA bands visible when the profile is visualized in gel electrophoresis with UV luminescence. This study will be a step and initial information about the genetic composition of a population of arabica coffee which still exists, and will be developed through DNA amplification technique. Key words: coffee, isolation of genomic DNA, CTAB method, profile band pattern

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Optimizing Genomic DNA Isolation and PCR Amplification For Pasak Bumi (Eurycoma longifolia)

KnE Engineering ◽

10.18502/keg.v1i2.4429 ◽

2019 ◽

Vol 1 (2) ◽

pp. 30

Author(s):

Arida Susilowati ◽

Henti Hendalastuti Rachmat ◽

Ahmad Baiquni Rangkuti ◽

Deni Elfiati ◽

Ami Ambarwati

Keyword(s):

Genomic Dna ◽

Dna Isolation ◽

Pcr Amplification ◽

Eurycoma Longifolia ◽

Genomic Dna Isolation

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Comparative Analysis of the Genomic DNA Isolation Methods on Inula sp. (Asteraceae)

Notulae Scientia Biologicae ◽

10.15835/nsb849881 ◽

2016 ◽

Vol 8 (4) ◽

pp. 451-455

Author(s):

Emre SEVİNDİK ◽

Fatih COŞKUN ◽

Zehra Tuğba MURATHAN ◽

Mehmet Yavuz PAKSOY ◽

Veysel UZUN

Keyword(s):

Genomic Dna ◽

Dna Isolation ◽

Low Cost ◽

Pcr Amplification ◽

Isoamyl Alcohol ◽

Dna Amount ◽

Genomic Dna Isolation ◽

Asteraceae Family ◽

Positive Results ◽

Commercial Kit

Simple, fast, low-cost and high throughput protocols are required for DNA isolation of plant species. In this study, phenol chloroform isoamyl alcohol and commercial (Sigma) DNA isolation kit methods were applied on some Inula species that belong to Asteraceae family. Genomic DNA amounts, A260, A280, A260/A230 and purity degrees (A260/A280) that were obtained through both methods were measured through electrophoresis and spectrophotometer. Additionally, PCR amplification was realized by primer pairs specific to nrDNA ITS, cpDNA ndhF (972F-1603R) and trnL-F regions. Results showed that maximum genomic DNA in nanograms obtained by phenol chloroform isoamyl alcohol method. The study also revealed that I. macrocephala had the maximum DNA and I. heterolepis had the minimum DNA amount. A260/A280 purity degrees showed that the highest and lowest purity in gDNAs obtained through phenol-choloform isoamyl alcohol method were in I.aucheriana and I. salicina, respectively. The highest and lowest purity degrees of gDNAs obtained through commercial kit was observed in I. fragilis and I. macrocephala samples, respectively. PCR amplification results showed that while band profiles of each three regions (ITS, trnL-F and ndhF) did not yield positive results in PCR amplifications using phenol-choloform isoamyl alcohol method; PCR band profiles obtained through commercial kit yielded positive results. As a result, it is fair to say that the relation of genomic DNA with PCR was found to be more efficient although the maximum amount of genomic DNA was obtained through phenol chloroform isoamyl alcohol method.

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Development of a Competent and Trouble Free DNA Isolation Protocol for Downstream Genetic Analyses in Glycine Species

Turkish Journal of Agriculture - Food Science and Technology ◽

10.24925/turjaf.v4i8.700-705.788 ◽

2016 ◽

Vol 4 (8) ◽

pp. 700 ◽

Cited By ~ 1

Author(s):

Muhammad Amjad Nawaz ◽

Faheem Shehzad Baloch ◽

Hafiz Mamoon Rehman ◽

Bao Le ◽

Fahima Akther ◽

...

Keyword(s):

Molecular Biology ◽

Dna Extraction ◽

Genomic Dna ◽

Dna Isolation ◽

Plant Molecular Biology ◽

Pcr Amplification ◽

Cost Effective ◽

Extraction Methods ◽

Glycine Species ◽

Genomic Dna Isolation

Extraction of deoxyribose nucleic acid (DNA) from plants is preliminary step in molecular biology. Fast and cost effective genomic DNA isolation from Glycine species for downstream application is a major bottleneck. Here we report a high throughput and trouble free method for genomic DNA extraction from leaf and seeds of Glycine species with high quality and quantity. Protocol reports the optimization by employing different concentrations of CTAB and PVP in extraction buffer. Efficiency of optimized protocol was compared with frequently used DNA extraction methods. Wide adoptability and utility of this protocol was confirmed by DNA extraction from leaves as well as seeds of G. max, G. soja, G. tomentella and G. latifolia. Extracted DNA was successfully subjected to PCR amplification of five microsatellite markers and four putative glycosyltransferase genes. DNA extraction protocol is reproducible, trouble free, rapid and can be adopted for plant molecular biology applications.

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