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

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

Journal of Applied and Advanced Research ◽

10.21839/jaar.2017.v2i4.97 ◽

2017 ◽

Vol 2 (4) ◽

Author(s):

S. Manikandan ◽

S. Ansarali ◽

G.M. Alagu Lakshmanan

Keyword(s):

Dna Barcoding ◽

Genomic Dna ◽

Dna Isolation ◽

Pcr Amplification ◽

Leaf Tissue ◽

Isolation Procedure ◽

Modified Method ◽

Genomic Dna Isolation ◽

Young Leaves ◽

Plectranthus Amboinicus

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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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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