IRESite record type: plasmid_with_promoter_and_putative_IRES_translationally_characterized
The shape of the nucleic acid molecule translated: linear
The quality of the mRNA/+RNA sequence: hopefully_full-length_mRNA
The mRNA/+RNA description:
In vitro T7 transcript containing renilla_luciferase - hairpin_from_multiple_cloning_site -
Kv1.4 IRES - firefly_luciferase and terminated at XbaI site immediately after the STOP codon. An FLuc-fusion
protein should be translated from the second cistron (4 aminoacids added extra).
The mRNA/+RNA sequence represented in the +DNA notation:
Credibility of mRNA sequence: end-to-end_sequence_reverse_engineered_and_should_match_experiment
The IRES absolute position (the range includes START and STOP codons or their equivalents): 1107-2304
How IRES boundaries were determined: experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF: 139
3'-end of IRES relative to last base of the STOP codon of the upstream ORF: 1336
5'-end of IRES relative to first base of the START codon of the downstream ORF: -1198
3'-end of IRES relative to first base of the START codon of the downstream ORF: -1
The sequence of IRES region aligned to its secondary structure (if available):
Remarks:
The results of Jang et al (2004) and of Jimenez et al (2005) disproved the former report of IRES
activity in KCNA4/Kv1.4 1.2, 1.0 and 0.2 kb segments by Negulescu et al. (1998).
The relative translation efficiency in % of this IRES: 9.100
Name of IRES used as the positive control: CVB3
Name of the plasmid used as the positive control. pRSTF-CVB3
Name of the plasmid used as the negative control. pRSTF
IRESite Id of the plasmid used as positive control. 210
IRESite Id of the plasmid used as negative control. 212
The relative translation efficiency in % of the positive control: 100.000
The relative translation efficiency in % of the negative control: 0.500
The size (length) of intercistronic region in the positive control: 819
The size (length) of intercistronic region in the negative control: 133
The effect of 5'-cap analogs on translation: not tested
Rapamycin affects translation: not tested
Type of RNA subject to translation: exogenous_RNA_without_cap_without_polyA_tail
Remarks:
The data are from Fig. 2B. Although Jang et al. (2004) did not state it explicitly the in vitro
transcripts used in the direct RNA transfection were most probably uncapped (this artificially increases the
yields from the second cistron and thus makes the IRES appear stronger and also increases the risk of degraded
RNA). In comparison with CVB3 IRES the Kv1.4 1.2kb fragment seriously underperforms. The transcripts should
have also been capped in yet another experiment. In the legend they mentioned measuring firefly
luciferase/beta-galactosidase measurements in these cells but that does not make sense as beta-galactosidase
was only measured in experiment shown in Fig. 2A.