An Optimised High-Salt CTAB Protocol for Both DNA and RNA Isolation from Succulent Stems of Hylocereus sp.

Abstract

The isolation of high quality DNA and RNA from plant species harboring high levels of polysaccharides and secondary metabolites are typically problematic, especially those in cactus. These compounds often co-precipitate with DNA and RNA thus causes low recovery and quality of the nucleic acids. Six DNA extraction protocols were tested on the sample of Hylocereus spp. of which the results were compared and analyzed. For comparison, three manufacturer’s protocols from different commercial kits and another three conventional DNA extraction protocols were compared. It was found that conventional method generally produces consistent and higher yield. Among the conventional protocols itself, each has their pros and cons. Therefore, a modified protocol which is concise, quick and simple was developed for Hylocereus spp. which is beneficial for further molecular work. This method was proven to be reliable in generating a good quality of DNA from these particular genera. Similarly for RNA extraction, four different extraction protocols were tested on the same sample. The results were analyzed and a modified protocol was developed to obtain a higher quality and yield of RNA for further downstream investigations. The extraction buffer from our modified DNA extraction protocol was tested on the RNA extraction and shows a good yield.