The nucleic acid data:
IRESite Id: 159 Version: 1
Originaly submitted by: Martin Mokrejš
Reviewed by: Martin Mokrejš Last change: 2007-11-02 00:00:00
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: 
Bicistronic mRNA transcribed by T7 polymerase in vitro or in vivo in BT7-H cells transiently expressing T7 Pol
coding for RLuc and CAT reporter genes separated by GBV-B IRES segment truncated by deletion of bases 136-153.
The mRNA/+RNA sequence represented in the +DNA notation:


Credibility of mRNA sequence:
  end-to-end_sequence_reverse_engineered_and_should_match_experiment
The name of the plasmid:
pBL-RLuc-GBB+3-delta-IIb-CATT
The name of the promoter used to express this mRNA:
  T7
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:
GBV-B
The origin of IRES in the plasmid:
  viral
The donor organism of the IRES segment:
Hepatitis GB virus B
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: pBL-RLuc-GBB+3-delta-IIb-CATT.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):
  32-967
ORF
ORF position:   2
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The abbreviated name of this ORF/gene:
CAT
The description of the protein encoded in this ORF:
Chloramphenicol acetyltransferase
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):
  1450-2121
Remarks:
The integrity of in vitro synthesized T7 transcripts was verified by agarose gel electrophoresis. The
integrity of in vivo transcripts synthesized in BT7-H cells was dependent on the Tpsi terminator function and was
not confirmed in this work.
Citations:
Rijnbrand R., Abell G., Lemon S. M. (2000) Mutational analysis of the GB virus B internal ribosome entry site. J. Virol. 74(2):773-783
IRESs:
IRES:
Version: 0
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  GBV-B-delta136-153
The functional status of IRES:
  defective
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  1023-1449
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  56
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  482
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -427
3'-end of IRES relative to first base of the START codon of the downstream ORF:
  -1
The sequence of IRES region aligned to its secondary structure (if available):


Citations:
Rijnbrand R., Abell G., Lemon S. M. (2000) Mutational analysis of the GB virus B internal ribosome entry site. J. Virol. 74(2):773-783
The translation experiments:
Translation results:
IRESite Translation Id: 117
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:
rabbit reticulocytes lysate
The organism used for translation:
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  43.000
Name of IRES used as the positive control:
  GBV-B
Name of the plasmid used as the positive control.
pBL-RLuc-GBB+3-CATT
IRESite Id of the plasmid used as positive control.
  153
The relative translation efficiency in % of the positive control:
  100.000
The size (length) of intercistronic region in the positive control:
500
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:
Fig. 3c
Citations:
Rijnbrand R., Abell G., Lemon S. M. (2000) Mutational analysis of the GB virus B internal ribosome entry site. J. Virol. 74(2):773-783
Translation results:
IRESite Translation Id: 118
Version: 2 Last change: 2006-07-13 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 BT7-H
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  15.000
Name of IRES used as the positive control:
  GBV-B
Name of the plasmid used as the positive control.
pBL-RLuc-GBB+3-CATT
IRESite Id of the plasmid used as positive control.
  153
The relative translation efficiency in % of the positive control:
  100.000
The size (length) of intercistronic region in the positive control:
500
The effect of 5'-cap analogs on translation:
not tested
Rapamycin affects translation:
not tested
Type of RNA subject to translation:
  endogenous_cytoplasmic_uncapped_T7_transcript_without_polyA_tail
Remarks:
Fig. 3d
Citations:
Rijnbrand R., Abell G., Lemon S. M. (2000) Mutational analysis of the GB virus B internal ribosome entry site. J. Virol. 74(2):773-783
Last change to the database: 2011-04-08 22:43:22 GMT+1