Comparative Different DNA Isolation Protocols from Ziziphus spina-christi (L.) Leaves through RAPD and ISSR Markers

Fatemeh Bina,Daryush Talei,Vahideh Nazeri,Zabihollah Zamani

doi:10.5539/jas.v8n6p49

Abstract

<p>Genomic analysis of plants relies on high quantity and quality of pure DNA. Extraction and purification of DNA from woody and medicinal plants, such as fruit trees present a great challenge due to accumulation of a large amount of co-purify with DNA, including polysaccharides, polyphenols and proteins. Therefore, it is necessary to optimize the extraction protocols to reduce these compounds to the lowest level. A study was conducted to compare six DNA extraction and precipitation methods for genomic analysis in<em> Ziziphus spina-christi</em> (L.) plant tissues. The results showed significant differences in DNA contents among the six methods. Quantity and quality of extracted genomic DNAs were compared by employing the spectrophotometer, Nano-Drop, agarose gel electrophoresis, digestion by restriction enzymes and polymerase chain reaction (PCR) methods and molecular marker such as RAPD and ISSR. The method of Vroh Bi et al., provided the best results (208.89 ng/μL) in terms of quantity and quality of DNA, and Doyle and Doyle method as second method for leaves sample were chosen. According to the results, the method of Bi et al. is recommended for DNA extraction from plant tissues having high level of polysaccharides and phenol compounds.</p>

Highlights

Ziziphus spina-christi (L.) is a tree indigenous to the south of Iran
Quantity and quality of extracted genomic DNAs were compared by employing the spectrophotometer, Nano-Drop, agarose gel electrophoresis, digestion by restriction enzymes and polymerase chain reaction (PCR) methods and molecular marker such as Random Amplified Polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR)
One gram of the frozen leaf samples of Z. spina-christi was ground into fine powder using pre-cooled mortar and pestle, and homogenized with six different DNA extraction methods based on randomized complete block design (RCBD) with five replicates

Summary

Introduction

Ziziphus spina-christi (L.) is a tree indigenous to the south of Iran. The leaves of this plant, which are locally known as “Sedr” and “Konar”, have been used for washing the hair and body (Anthony, 2005). Aqueous leaf extract of Z. spina-christi may possess anti-nociceptive properties in the rat and have a calming effect on the central nervous system (Effraim et al, 1998). It has been described as ant cathartic, astringent, diuretic and tonic (Larsen, 1985). The species is ecologically and economically important due to fruit nutrition, leaves and shoots saponin, and tannin substrates (Sudhersan & Hussain, 2003; Weinges & Schick, 1995)

Methods

Results

Conclusion