The nucleic acid data:
IRESite Id: 605 Version: 3
Originaly submitted by: Martin Mokrejš Submission date: 2009-08-23 18:08:46
Reviewed by: Martin Mokrejš Last change: 2009-08-26 13:44:05
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:
  both_UTRs_incomplete
The mRNA/+RNA description: 
Part of the T7-derived transcript studied in in vitro assays. The 5'-UTR could have been longer but do not
know the necessary details. The intercistronic region contains IRES sequence of crTMV virus (bp -149 to -1
relative to coat protein ORF).
The mRNA/+RNA sequence represented in the +DNA notation:


Warning: available plasmid sequence is exactly same as the mRNA sequence. It seems we never managed to reconstruct the original plasmid sequence used to express the studied mRNA.
Credibility of mRNA sequence:
  reverse_engineered_fragment_and_the_rest_is_a_guess
The name of the plasmid:
H-CP-IREScp-GUS
The name of the promoter used to express this mRNA:
  T7
Aliases of the plasmid name:
Alias: hCP-IREScp-GUS
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:
crTMV
The origin of IRES in the plasmid:
  viral
The donor organism of the IRES segment:
Crucifer tobamovirus
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:
H-CP-IREScp-GUS.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:
CP
The description of the protein encoded in this ORF:
coat protein
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):
  77-550
ORF
ORF position:   2
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
GUS
The description of the protein encoded in this ORF:
beta-glucuronidase
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):
  730-2541
Remarks:
A single-read sequencing result between CP and GUS ORFs obtained from T. Komarova.
Citations:
Ivanov P. A., Karpova O. V., Skulachev M. V., Tomashevskaya O. L., Rodionova N. P., Dorokhov YuL, Atabekov J. G. (1997) A tobamovirus genome that contains an internal ribosome entry site functional in vitro. Virology. 232(1):32-43
IRESs:
IRES:
Version: 2 Last change: 2009-08-26 13:44:05
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  crTMV_IREScp
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  580-727
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  30
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  177
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -150
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:
Ivanov P. A., Karpova O. V., Skulachev M. V., Tomashevskaya O. L., Rodionova N. P., Dorokhov YuL, Atabekov J. G. (1997) A tobamovirus genome that contains an internal ribosome entry site functional in vitro. Virology. 232(1):32-43
The translation experiments:
Translation results:
IRESite Translation Id: 632
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vitro
The in vitro translation system:
wheat germ lysate
The organism used for translation:
The temperature (in degrees of Celsia):
30
The relative translation efficiency in % of this IRES:
  100.000
Name of the plasmid used as the negative control.
H-CP-TMV_UI-GUS
IRESite Id of the plasmid used as negative control.
  604
The relative translation efficiency in % of the negative control:
  0
The size (length) of intercistronic region in the negative control:
80
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:
Data reflect results shown in Figure 7B. However, IRESite did not quantify intensities of the bands.
Citations:
Ivanov P. A., Karpova O. V., Skulachev M. V., Tomashevskaya O. L., Rodionova N. P., Dorokhov YuL, Atabekov J. G. (1997) A tobamovirus genome that contains an internal ribosome entry site functional in vitro. Virology. 232(1):32-43
Translation results:
IRESite Translation Id: 633
Version: 1 Last change: 2009-08-23 20:26:04
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The translation method used to study IRES function:
in vitro
The in vitro translation system:
rabbit reticulocytes lysate
The organism used for translation:
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.
H-CP-TMV_UI-GUS
IRESite Id of the plasmid used as negative control.
  604
The relative translation efficiency in % of the negative control:
  0
The size (length) of intercistronic region in the negative control:
80
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:
Data reflect results shown in Figure 7B. However, IRESite did not quantify intensities of the bands.

Only GUS protein is produced from this message. Removal of the hairpin from the 5'-end (CP-IREScp-GUS plasmid)
results in synthesis of the CP protein while almost disabling the IRES-mediated synthesis of GUS.
Citations:
Ivanov P. A., Karpova O. V., Skulachev M. V., Tomashevskaya O. L., Rodionova N. P., Dorokhov YuL, Atabekov J. G. (1997) A tobamovirus genome that contains an internal ribosome entry site functional in vitro. Virology. 232(1):32-43
Last change to the database: 2015-04-16 16:45:23 GMT+1