The nucleic acid data:
IRESite Id: 355 Version: 1
Originaly submitted by: Martin Mokrejš
Reviewed by: Václav Vopálenský Last change: 2008-06-27 12:39:37
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: 
Putative spliced transcript of phpRMF plasmid from SV40 promoter containing full-length 5'-UTR (1-395 bp) of
c-myc mRNA transcribed from major P2 promoter inserted between renilla and firefly luciferases as PvuII-NcoI
insert with a hairpin in front of the first cistron
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:
  end-to-end_sequence_reverse_engineered_and_should_match_experiment
The name of the plasmid:
phpRMF
The name of the promoter used to express this mRNA:
  SV40
Aliases of the plasmid name:
Alias: pGL3RutrH
Alias: pstemRmycF
Alias: pGL3R2utr
The in vivo produced transcripts are heterogeneous (due to any of promoter?/splicing?/cleavage?/breakage?):
  not tested
The in vivo produced heterogeneous transcripts occur due to alternative splicing:
  not tested
A promoter reported in cDNA corresponding to IRES sequence:
  not tested
The abbreviated name of the donor gene or virus from which this IRES was excised and inserted into the plasmid:
c-myc
The origin of IRES in the plasmid:
  nuclear
The donor organism of the IRES segment:
Homo sapiens
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:
phpRMF.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: 2008-06-27 12:39:37
Originaly submitted by: Martin Mokrejš Reviewed by: Václav Vopálenský
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):
  246-1181
ORF
ORF position:   2
Version: 1 Last change: 2008-06-27 12:39:37
Originaly submitted by: Martin Mokrejš Reviewed by: Václav Vopálenský
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):
  1628-3280
Citations:
Stoneley M., Paulin F. E., Le Quesne J. P., Chappell S. A., Willis A. E. (1998) C-Myc 5' untranslated region contains an internal ribosome entry segment. Oncogene. 16(3):423-428
IRESs:
IRES:
Version: 1 Last change: 2008-06-27 12:39:37
Originaly submitted by: Martin Mokrejš Reviewed by: Václav Vopálenský
The IRES name:
  c-myc_1-395
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  1232-1625
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  51
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  444
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -396
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:
Stoneley M., Paulin F. E., Le Quesne J. P., Chappell S. A., Willis A. E. (1998) C-Myc 5' untranslated region contains an internal ribosome entry segment. Oncogene. 16(3):423-428
The translation experiments:
Translation results:
IRESite Translation Id: 428
Version: 3 Last change: 2008-07-08 17:54:46
Originaly submitted by: Martin Mokrejš Reviewed by: Václav Vopálenský
The translation method used to study IRES function:
in vivo
The organism used for translation:
Homo sapiens HeLa (ATCC CCL-2)
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  387.000
Name of IRES used as the positive control:
  c-myc_1-395
Name of the plasmid used as the positive control.
pRMF
IRESite Id of the plasmid used as positive control.
  25
The relative translation efficiency in % of the positive control:
  100.000
The size (length) of intercistronic region in the positive control:
446
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:
As a positive control was used HeLa cell line transformed by similar vector pRMF lacking the stem-loop in
front of of the first cistron. The hairpin loop impairs translation (by 75%) from the first cistron while
translation from the second cistron is not affected. Thus, the relative ratio 2nd/1st citron increases. Data
from Fig. 2c. The original relative renilla/firefly luciferase activities were normalized by authors against
beta-galactosidase activity (co-transfection control).
Citations:
Stoneley M., Paulin F. E., Le Quesne J. P., Chappell S. A., Willis A. E. (1998) C-Myc 5' untranslated region contains an internal ribosome entry segment. Oncogene. 16(3):423-428
Translation results:
IRESite Translation Id: 518
Version: 1 Last change: 2008-07-08 18:16:05
Originaly submitted by: Václav Vopálenský Reviewed by: Václav Vopálenský
The translation method used to study IRES function:
in vivo
The organism used for translation:
Homo sapiens HeLa (ATCC CCL-2)
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  100.000
Name of the plasmid used as the negative control.
pstemRF
IRESite Id of the plasmid used as negative control.
  455
The relative translation efficiency in % of the negative control:
  0
The size (length) of intercistronic region in the negative control:
40
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:
Translational data are derived from Fig. 5B.
Citations:
Lang K. J. D., Kappel A., Goodall G. J. (2002) Hypoxia-inducible factor-1alpha mRNA contains an internal ribosome entry site that allows efficient translation during normoxia and hypoxia. Mol. Biol. Cell. 13(5):1792-1801
Translation results:
IRESite Translation Id: 519
Version: 1 Last change: 2008-07-08 18:16:05
Originaly submitted by: Václav Vopálenský Reviewed by: Václav Vopálenský
The translation method used to study IRES function:
in vivo
The organism used for translation:
Homo sapiens NIH/3T3 (ATCC CRL-1658)
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  100.000
Name of the plasmid used as the negative control.
pstemRF
IRESite Id of the plasmid used as negative control.
  455
The relative translation efficiency in % of the negative control:
  0
The size (length) of intercistronic region in the negative control:
40
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:
Translational data are derived from Fig. 5A.
Citations:
Lang K. J. D., Kappel A., Goodall G. J. (2002) Hypoxia-inducible factor-1alpha mRNA contains an internal ribosome entry site that allows efficient translation during normoxia and hypoxia. Mol. Biol. Cell. 13(5):1792-1801
Last change to the database: 2015-04-16 16:45:23 GMT+1