pELMO, an optimised in-house cloning vector Academic Article

abstract

  • 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.

publication date

  • 2017/12/1

edition

  • 7

keywords

  • Costs and Cost Analysis
  • DNA
  • DNA Replication
  • Enzymes
  • Genes
  • Genetic Vectors
  • Molecular Biology
  • Organism Cloning
  • Plasmids
  • Polymerase Chain Reaction
  • Recombinant DNA
  • Technology

International Standard Serial Number (ISSN)

  • 2191-0855