Optimization of DNA extraction protocol for leaf of narcotic plant Cannabis sativa L.
Muhammad Ishtiaq, Muhammad Siddique, Shamaila Nazeer, Waheeda Mushtaq
Cannabis sativa L. (CSL), it is very important plant being for industrial and medicinal purposes. All of its three very important species (Cannabis sativa, Cannabis indica, Cannabis ruderalis) highly prohibited to their cultivate, so it makes very hard for those who want to conduct studies on genetic and molecular properties of plant. More DNA is the only material does not change in response to environmental and other changes and contains all the answers of mysteries. So studies are conducted on the genomic DNA of plant. So highly efficient DNA extraction method is required because plants tissues are not available easily. We have tested four different protocols and became able to give the best optimized DNA extraction protocol (of yield 168 μg/g). Our optimized protocol replaces use of liquid nitrogen which is very expensive by alcohol fixing the leaves in the fridge for four days before use and washing with phenol: chloroform: isoamyl: alcohol, Polyvinylpyrrolidone and beta-mercaptoethanol which gave high quantity of the DNA per leaf by knocking out Secondary metabolites (polyphenols and polysaccharides) which cause contamination of genomic DNA by forming a layer like mist. It gave successful amplification of genes and shows bands of very high quality without degradation and contamination on the Agarose gel electrophoresis.