Improved System for Constructing Bacterial cDNA Libraries From the Venom Glands of Two Iranian Scorpions

Abstract

Background: Analyzing a cell or a tissue transcriptome is the best way to understand its functions. cDNA library construction is one of the best methods to achieve this aim, and it is used to determine the structure, functions, and expression profile of a transcriptome. In the construction of a cDNA library, bacterial hosts are very adaptive. One of the most adaptive bacterial hosts is the DH5α strain of Escherichia coli. Objectives: The venom peptides of scorpions are applicable in biological and medical research as potent drugs, as they have bioactive components; therefore, their identification is very important. In this project, we set up and improved the Clontech® In-Fusion SMARTer™ Directional cDNA library construction kit in the transcriptome analysis of the venom glands of two Iranian scorpions, Odonthubuthus doriae (O. doriae) and Mesobuthus eupeus (M. eupeus).Methods: RNA extraction and cDNA synthesis and purification were conducted on the venom glands of O. doriae and M. eupeus scorpions. cDNA libraries were constructed by cloning and transforming chemically competent bacterial cells. The blue-white screen method was used to identify the transformed cells. For finding colonies that have scorpion-related inserts, a colony PCR method was designed with specific primers for two sides of the insertion site into the vector. Results: The obtained white colonies have vectors that include cDNA related to O. doriae and M. eupeus. Conclusions:The constructed cDNA libraries of O. doriae and M. eupeus glands confirmed that a framework was created for the transcriptome analysis and peptide study of the venom glands of other scorpion species.