The nucleic acid data:
IRESite Id: 228 Version: 5
Originaly submitted by: Martin Mokrejš
Reviewed by: Martin Mokrejš Last change: 2008-06-10 17:08:56
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:
  our_best_guess
The mRNA/+RNA description: 
Putative spliced bicistronic mRNA derived from pRF based plasmid with HRV IRES in the intercistronic region.
The mRNA/+RNA sequence represented in the +DNA notation:


Warning: mRNA sequence when devoid of trailing 'A's is still not a substring of the plasmid sequence. Is it because an intron is spliced out? Stay calm then. :-)
Credibility of mRNA sequence:
  reverse_engineered_sequence_and_should_match_experiment_maybe_except_both_UTRs
The name of the plasmid:
pR-HRV-F
The name of the promoter used to express this mRNA:
  SV40
The in vivo produced transcripts are heterogeneous (due to any of promoter?/splicing?/cleavage?/breakage?):
  no
The in vivo produced heterogeneous transcripts occur due to alternative splicing:
  no
A promoter reported in cDNA corresponding to IRES sequence:
  no
Summary of possible issues when IRES cDNA is experimentally transcribed in vivo:
Summary of experiments studying integrity of the in vivo transcripts in a particular host:
Integrity (uniformity) of mRNA tested using Northern-blot:
homogeneous_population_of_molecules_confirmed
Integrity (uniformity) of mRNA tested using RNase protection:
not_tested
Integrity (uniformity) of mRNA tested using 5'-RACE:
not_tested
Integrity (uniformity) of mRNA tested using primer extension :
not_tested
Integrity (uniformity) of mRNA tested using RT-PCR:
not_tested
Integrity (uniformity) of mRNA tested using real-time quantitative polymerase chain reaction (rtqPCR):
not_tested
Integrity (uniformity) of mRNA tested using RNAi:
not_tested
Integrity (uniformity) of mRNA tested using S1 nuclease mapping:
not_tested
Cryptic promoter presence was confirmed by expression from a promoter-less plasmid:
no_promoter_confirmed
Cryptic promoter presence was confirmed in an experimental setup involving inducible promoter:
not_tested
Integrity (uniformity) of mRNA molecules or possible promoter presence expressed in vivo was tested using another method, please specify in Remarks:
not_tested
The organism used:
Cercopithecus aethiops COS-7 (ATCC CRL-1651)
The abbreviated name of the donor gene or virus from which this IRES was excised and inserted into the plasmid:
HRV2
The origin of IRES in the plasmid:
  viral
The donor organism of the IRES segment:
Human rhinovirus 2
The DNA sequence of the plasmid in (+) orientation annotated by its secondary structure:


GenBank formatted file with annotated plasmid sequence hyperlinked from vector image map: pR-HRV-F.jpg
The total number of notable open-reading frames (ORFs):
  2
Notable Open-Reading Frames (ORFs; protein coding regions) in the mRNA/+RNA sequence:
ORF
ORF position:   1
Version: 1 Last change: 2008-05-22 19:09:57
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
RLuc
The description of the protein encoded in this ORF:
Renilla luciferase
The translational frameshift (ribosome slippage) involved:
  0
The ribosome read-through involved:
  no
The alternative forms of this protein occur by the alternative initiation of translation:
  not tested
The ORF absolute position (the base range includes START and STOP codons or their equivalents):
  192-1127
ORF
ORF position:   2
Version: 1 Last change: 2008-05-22 19:09:57
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
FLuc
The description of the protein encoded in this ORF:
Firefly luciferase
The translational frameshift (ribosome slippage) involved:
  0
The ribosome read-through involved:
  no
The alternative forms of this protein occur by the alternative initiation of translation:
  not tested
The ORF absolute position (the base range includes START and STOP codons or their equivalents):
  1784-3436
Remarks:
The plasmid with SpeI - NcoI insert has been created by Han and Zhang (2002) and the sequence has been
provided to us by Wang et al. (2005). Han and Zhang (2002) re-cloned the PvuII - NcoI HRV-2 insert of
Stoneley et al. (2000) as SpeI - NcoI into the same backbone vector pRF. It is not clear why they did
that.
No additional transcripts were found on Northern-blot by Wang et al. from pR-HRV-F plasmid as shown in
Fig. 6A and B.
Citations:
Wang Z., Weaver M., Magnuson N. S. (2005) Cryptic promoter activity in the DNA sequence corresponding to the pim-1 5'-UTR. Nucleic Acids Res. 33(7):2248-2258
Han B., Zhang J. T. (2002) Regulation of gene expression by internal ribosome entry sites or cryptic promoters: the eIF4G story. Mol. Cell. Biol. 22(21):7372-7384
IRESs:
IRES:
Version: 4 Last change: 2008-05-22 19:09:57
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  HRV-2
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  1168-1781
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  41
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  654
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -616
3'-end of IRES relative to first base of the START codon of the downstream ORF:
  -3
The sequence of IRES region aligned to its secondary structure (if available):


Citations:
Wang Z., Weaver M., Magnuson N. S. (2005) Cryptic promoter activity in the DNA sequence corresponding to the pim-1 5'-UTR. Nucleic Acids Res. 33(7):2248-2258
The translation experiments:
Translation results:
IRESite Translation Id: 293
Version: 1 Last change: 2006-09-25 00:00:00
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vivo
The organism used for translation:
Cercopithecus aethiops COS-7 (ATCC CRL-1651)
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  110.900
Name of IRES used as the positive control:
  EMCV
Name of the plasmid used as the positive control.
pR-EMCV-F
Name of the plasmid used as the negative control.
pRF
IRESite Id of the plasmid used as positive control.
  227
IRESite Id of the plasmid used as negative control.
  184
The relative translation efficiency in % of the positive control:
  100.000
The relative translation efficiency in % of the negative control:
  8.100
The size (length) of intercistronic region in the positive control:
632
The size (length) of intercistronic region in the negative control:
67
The effect of 5'-cap analogs on translation:
not tested
Rapamycin affects translation:
not tested
Type of RNA subject to translation:
  endogenous_nuclear_RNA_Pol_II_transcript
Remarks:
Fig. 2b
Citations:
Wang Z., Weaver M., Magnuson N. S. (2005) Cryptic promoter activity in the DNA sequence corresponding to the pim-1 5'-UTR. Nucleic Acids Res. 33(7):2248-2258
Last change to the database: 2011-04-08 22:43:22 GMT+1