The nucleic acid data:
IRESite Id: 404 Version: 1
Originaly submitted by: Václav Vopálenský
Reviewed by: Martin Mokrejš Last change: 2008-06-20 12:58:28
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:
  end-to-end_full-length_mRNA
The mRNA/+RNA description: 
In vitro T3 run-off transcript containing 5' UTR of human c-myc (nt from -407 to -2) mRNA transcribed from
major P2 promoter cloned upstream of the luciferase reporter RNA lacking a poly (A) tail.
The mRNA/+RNA sequence represented in the +DNA notation:


Credibility of mRNA sequence:
  end-to-end_sequence_completely_same_as_in_the_experiment
The name of the plasmid:
pML
The name of the promoter used to express this mRNA:
  T3
Description of the plasmid (facultative for promoter-less plasmid records):
Plasmid containing 5' UTR of human c-myc transcribed from major P2 promoter (nt from -407 to -2) and firefly luciferase coding region (luc version of the luciferase gene) without the poly(A)tract.
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 HeLa S3 (ATCC CCL-2.2)
The DNA sequence of the plasmid in (+) orientation annotated by its secondary structure:


Plasmid sequence verified (partially/completely) by IRESite (more details in Remarks):
  plasmid sequence confirmed by IRESite curators by restriction analysis + parts by PCR + sequencing
GenBank formatted file with annotated plasmid sequence hyperlinked from vector image map:
pML.jpg
The total number of notable open-reading frames (ORFs):
  1
Notable Open-Reading Frames (ORFs; protein coding regions) in the mRNA/+RNA sequence:
ORF
ORF position:   1
Version: 0
Originaly submitted by: Václav Vopálenský Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
FFluc
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):
  500-2155
Remarks:
IRESite notes about verification of plasmid sequence:
The complete sequence of IRES element was determined by sequencing using Fluc_seq_reverse
(AGGAACCAGGGCGTATCTC) primer.

For detailed verification of whole plasmid sequence see item IRESite Id: 413.
Citations:
Thoma C., Bergamini G., Galy B., Hundsdoerfer P., Hentze M. W. (2004) Enhancement of IRES-mediated translation of the c-myc and BiP mRNAs by the poly(A) tail is independent of intact eIF4G and PABP. Mol. Cell. 15(6):925-935
IRESs:
IRES:
Version: 1 Last change: 2008-06-19 14:18:19
Originaly submitted by: Václav Vopálenský Reviewed by: Martin Mokrejš
The IRES name:
  c-myc
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  92-498
How IRES boundaries were determined:
experimentally_determined
The sequence of IRES region aligned to its secondary structure (if available):


Citations:
Thoma C., Bergamini G., Galy B., Hundsdoerfer P., Hentze M. W. (2004) Enhancement of IRES-mediated translation of the c-myc and BiP mRNAs by the poly(A) tail is independent of intact eIF4G and PABP. Mol. Cell. 15(6):925-935
The translation experiments:
Translation results:
IRESite Translation Id: 461
Version: 3 Last change: 2008-06-20 12:58:28
Originaly submitted by: Václav Vopálenský Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vitro
The in vitro translation system:
HeLa cell lysate
The organism used for translation:
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  31.500
Name of IRES used as the positive control:
  c-myc
Name of the plasmid used as the positive control.
pML-p(A)
Name of the plasmid used as the negative control.
pMiL
IRESite Id of the plasmid used as positive control.
  403
IRESite Id of the plasmid used as negative control.
  399
The relative translation efficiency in % of the positive control:
  100.000
The relative translation efficiency in % of the negative control:
  7.800
The effect of 5'-cap analogs on translation:
no
Rapamycin affects translation:
not tested
Type of RNA subject to translation:
  exogenous_RNA_with_ApppG_cap_without_polyA_tail
Remarks:
The translation of c-myc.IRES RNA is about 3-fold less efficient than the translation of the polyadenylated
counterpart c-myc.IRES-pA RNA (IRESite ID: 399).

Translational data are derived from Fig. 1A.
Citations:
Thoma C., Bergamini G., Galy B., Hundsdoerfer P., Hentze M. W. (2004) Enhancement of IRES-mediated translation of the c-myc and BiP mRNAs by the poly(A) tail is independent of intact eIF4G and PABP. Mol. Cell. 15(6):925-935
Translation results:
IRESite Translation Id: 462
Version: 1 Last change: 2008-06-19 10:54:01
Originaly submitted by: Václav Vopálenský Reviewed by: Martin Mokrejš
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:
  0
Name of IRES used as the positive control:
  c-myc
Name of the plasmid used as the positive control.
pMiL-p(A)
Name of the plasmid used as the negative control.
pNO-IRES
IRESite Id of the plasmid used as positive control.
  403
IRESite Id of the plasmid used as negative control.
  394
The relative translation efficiency in % of the positive control:
  100.000
The relative translation efficiency in % of the negative control:
  0
The effect of 5'-cap analogs on translation:
not tested
Rapamycin affects translation:
not tested
Type of RNA subject to translation:
  exogenous_RNA_with_ApppG_cap_without_polyA_tail
Remarks:
The poly(A) tail is essential for cellular IRES-driven translation in transfected cells.
RNAs lacking a poly(A) tail are poorly translated, only the presence of a poly(A) tail essentially enables
translation (see IRESId: 403)

Translational data are derived from Fig. 2A.
Citations:
Thoma C., Bergamini G., Galy B., Hundsdoerfer P., Hentze M. W. (2004) Enhancement of IRES-mediated translation of the c-myc and BiP mRNAs by the poly(A) tail is independent of intact eIF4G and PABP. Mol. Cell. 15(6):925-935
Last change to the database: 2019-03-18 09:32:49 GMT+1