Size (kb): 6.2699999809265140
Vector: pIR2 (plasmid)
Construction: pDS, IRES
Construct size (kb): 6.269999980926514
Features: marker(s): ampR
other: 3' LTR
other: internal ribosomal entry site (IRES)
restriction site: SalI
coding sequence: 3' env
retroviral expression vector
Restriction digests of the clone give the following sizes (kb): EcoRV - 6.3; SalI - 3.7, 2.7; HindIII - 5.2, 1.1.
Avian retroviral expression vector containing an internal ribosomal entry site (IRES) for higher level expression, derived from Rous sarcoma virus (RSV), requiring vector pRep(A) (ATCC 87702)
for recombinant virus production.
Replication-competent viral vectors expressing foreign DNA can be recovered from the culture fluid of transfected cells after the following treatment:
Foreign DNA is cloned into the multiple cloning site, followed by digestion with SalI, ligation to SalI digested pRep(A) DNA and transfection into chicken primary cells or other appropriate cell lines.
The IRES, derived from encephalomyocarditis virus (EMCV), allows ribosomal binding and transcription within an mRNA transcript and increases expression levels 3 - 8 fold over the corresponding expression vector lacking IRES.
Vector lacks the splice acceptor site, found downstream of env in vector pIR1 (ATCC 87700)
. The lack of splice acceptor is expected to prevent production of the shortest subgenomic mRNA and ensures expression of the exogenous gene only from IRES.
Murakami M, et al. High-level expression of exogenous genes by replication-competent retrovirus vectors with an internal ribosomal entry site. Gene 202: 23-29, 1997. PubMed: 9427541