A Rapid and Efficient Method for the Isolation of Mitochondrial DNA From Wheat Crop

Abstract

The mitochondrial DNA was isolated from wheat (Triticum aestivum) crop in combination of different centrifugation and density gradient ultracentrifugation, DNaseI enzyme treatment, sucrose sedimentation; lysis with SDS and potassium proteinase. Pure mitochondrial DNA was successfully obtained by phenol/chloroform/isoamyl alcohol extraction to remove protein, and digested with RNase phenol/chloroform extraction method. To detect mitochondrial DNA purity, specific primers were designed for nuclear (β-actin) and mitochondrial (COXII) gene. Isolated mitochondrial DNA is pure, suitable for PCR and genetic analysis.

A Rapid and Efficient Method for the Isolation of Mitochondrial Dna From Small Amounts of Some Fruit Trees

“…Numerous mitochondrial DNA isolation protocols have been described in the literature (Ejaz et al, 2013, andHwang et al, 2001), all of which aim to maximize the mitochondrial DNA yield, while minimizing the contamination of nuclear DNA.…”

“…Phylogenetic and evolutionary relationships, (Schuster & Brennicke, 1994). Furthermore, isolation of plant mitochondrial DNA for use in different PCR techniques is one of the most important and time-consuming steps (Ejaz et al, 2013). Such techniques include the Southern blot analysis, polymerase chain reaction amplifications (AFLP, RAPD, SSR).…”

“…(Pérez et al, 1990;Keech et al, 2005;Cui et al, 2009) but most of them are either time consuming or ineffective when applied to small amounts of material. Procedures to isolate pure mitochondrial DNA are often drafted on two basic methodologies involving Percoll and Sucrose density gradient centrifugation (Keech et al, 2005;Cui et al, 2009;Ejaz et al, 2013). However, these methodologies often require an expensive laboratory setup which is not possible for many researchers in low-budget labs.…”