Mutagenesis to Replace IRES with T2A in NSD3-Short-3xFLAG Lentiviral Expression Vector

SGC Open Notebook Project to Characterize the HMTase NSD3

Exp016 Objective: Bicistronic elements are common features in lentiviral expression systems that allow expression of two protein products from a single promoter. The two most common bicistronic elements are the internal ribosome entry site (IRES), which allows cap-independent translation of a second open reading frame within an mRNA, and the 2A system, which introduces a self-cleaving peptide between two desired protein products. There are several advantages to the the 2A system over an IRES, which include matched expression levels of your two protein products of interest as well as having a smaller footprint (~1/10 size in kb), which may improve viral titre. Therefore, I will use site-directed mutagenesis to alter the NSD3short lentiviral expression plasmid to replace the IRES sequence with a T2A sequence.