The nucleic acid data:
IRESite Id: 585 Version: 0
Originaly submitted by: Martin Mokrejš Submission date: 2009-04-14 14:59:16
Reviewed by: Martin Mokrejš
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: 
In vivo spliced transcript derived through CMV IE promoter containing Renilla and firefly luciferases and
terminated after the SV40 late poly(A) signal.
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:
pR-EMCV-F
The name of the promoter used to express this mRNA:
  CMV_IE
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:
EMCV-R
The origin of IRES in the plasmid:
  viral
The donor organism of the IRES segment:
Encephalomyocarditis virus Rueckert
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-EMCV-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: 0
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):
  219-1154
ORF
ORF position:   2
Version: 0
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):
  1924-3576
Citations:
Petz M., Kozina D., Huber H., Siwiec T., Seipelt J., Sommergruber W., Mikulits W. (2007) The leader region of Laminin B1 mRNA confers cap-independent translation. Nucleic Acids Res. 35(8):2473-2482
IRESs:
IRES:
Version: 1 Last change: 2011-04-08 21:47:14
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  EMCV-R_260-831
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  1250-1829
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  96
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  675
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -674
3'-end of IRES relative to first base of the START codon of the downstream ORF:
  -95
The sequence of IRES region aligned to its secondary structure (if available):


Remarks:
The initial 7 nt do not align to the EMCV-R genome.
Citations:
Petz M., Kozina D., Huber H., Siwiec T., Seipelt J., Sommergruber W., Mikulits W. (2007) The leader region of Laminin B1 mRNA confers cap-independent translation. Nucleic Acids Res. 35(8):2473-2482
The translation experiments:
Translation results:
IRESite Translation Id: 619
Version: 2 Last change: 2009-04-30 11:34:57
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vivo
The organism used for translation:
Mus musculus MIM-R
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.
pR-F
IRESite Id of the plasmid used as negative control.
  584
The relative translation efficiency in % of the negative control:
  25.000
The size (length) of intercistronic region in the negative control:
112
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:
Measured luciferase activity values were provided by M. Petz for Figure 3C.

Additional notes from M Petz:

Concerning Fig. 3A, I have checked the data again. The experiment was repeated several times and the ß-Gal
activities of the cotransfected plamid were always quite equal. The normalisation to the ß-gal values did
not affect the ratio of the firefly activities in any significant way.
Citations:
Petz M., Kozina D., Huber H., Siwiec T., Seipelt J., Sommergruber W., Mikulits W. (2007) The leader region of Laminin B1 mRNA confers cap-independent translation. Nucleic Acids Res. 35(8):2473-2482
Translation results:
IRESite Translation Id: 620
Version: 2 Last change: 2011-04-08 21:47:14
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vivo
The organism used for translation:
Homo sapiens SW480
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.
pR-F
IRESite Id of the plasmid used as negative control.
  584
The relative translation efficiency in % of the negative control:
  18.000
The size (length) of intercistronic region in the negative control:
112
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:
Measured luciferase activity values were provided by M. Petz for Figure 3D.

Additional notes from M Petz:

Concerning Fig. 3A, I have checked the data again. The experiment was repeated several times and the ß-Gal
activities of the co-transfected plasmid were always quite equal. The normalization to the ß-gal values did
not affect the ratio of the firefly activities in any significant way.
Citations:
Petz M., Kozina D., Huber H., Siwiec T., Seipelt J., Sommergruber W., Mikulits W. (2007) The leader region of Laminin B1 mRNA confers cap-independent translation. Nucleic Acids Res. 35(8):2473-2482
Last change to the database: 2015-04-16 16:45:23 GMT+1