pELMO, an optimised in-house cloning vector

DNA cloning is an essential tool regarding DNA recombinant technology as it allows the replication of foreign DNA fragments within a cell. pELMO was here constructed as an in-house cloning vector for rapid and low-cost PCR product propagation; it is an optimally designed vector containing the ccdB killer gene from the pDONR 221 plasmid, cloned into the pUC18 vector’s multiple cloning site (Thermo Scientific). The ccdB killer gene has a cleavage site (CCC/GGG) for the SmaI restriction enzyme which is used for vector linearisation and cloning blunt-ended products. pELMO transformation efficiency was evaluated with different sized inserts and its cloning efficiency was compared to that of the pGEM-T Easy commercial vector. The highest pELMO transformation efficiency was observed for ~500 bp DNA fragments; pELMO vector had higher cloning efficiency for all insert sizes tested. In-house and commercial vector cloned insert reads after sequencing were similar thus highlighting that sequencing primers were designed and localised appropriately. pELMO is thus proposed as a practical alternative for in-house cloning of PCR products in molecular biology laboratories. © 2017, The Author(s).

• Fecha de publicación: 2017
• Institución: Universidad del Rosario
• Repositorio: Repositorio EdocUR - U. Rosario
• Idioma: eng
• OAI Identifier: oai:repository.urosario.edu.co:10336/18824
• Acceso en línea: http://repository.urosario.edu.co/handle/10336/18824
• Palabra clave: Blunt-Ended; Ccdb Killer Gene; Cloning Vector; Pcr Cloning; Recombinant Dna Technology; Plasmid Vector; Restriction Endonuclease; Article; Bacterium Transformation; Cloning Vector; Molecular Cloning; Neospora Caninum; Nonhuman; Pelmo Vector; Plasmodium Falciparum; Plasmodium Vivax; Polymerase Chain Reaction; Process Design; Process Optimization; Protein Cleavage; Sanger Sequencing
• License: Abierto (Texto completo)