The nucleic acid data:
IRESite Id: 261 Version: 1
Originaly submitted by: Martin Mokrejš
Reviewed by: Martin Mokrejš Last change: 2009-08-30 23:45:42
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:
  3UTR_possibly_incomplete
The mRNA/+RNA description: 
Putative bicistronic mRNA containing renilla and firefly luciferase genes with putative c-IAP1 IRES in the
intercistronic region without the chimeric intron. The sequence ends at its 3'-end right after the poly(A)
signal from bovine growth hormone (BGH) mRNA and thus the 3'-UTR might be slightly wrong (it is unknown
what region of the BGH signal could was transcribed and where the poly(A) tail was added).
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:
  reverse_engineered_fragment_and_the_rest_is_a_guess
The name of the plasmid:
pRL-cIAP1-FL
The name of the promoter used to express this mRNA:
  CMV
The in vivo produced transcripts are heterogeneous (due to any of promoter?/splicing?/cleavage?/breakage?):
  yes
The in vivo produced heterogeneous transcripts occur due to alternative splicing:
  yes
A promoter reported in cDNA corresponding to IRES sequence:
  no
Summary of possible issues when IRES cDNA is experimentally transcribed in vivo:
Summary of experiments studying integrity of the in vivo transcripts in a particular host:
Integrity (uniformity) of mRNA tested using Northern-blot:
not_tested
Integrity (uniformity) of mRNA tested using RNase protection:
not_tested
Integrity (uniformity) of mRNA tested using 5'-RACE:
not_tested
Integrity (uniformity) of mRNA tested using primer extension :
not_tested
Integrity (uniformity) of mRNA tested using RT-PCR:
heterogeneous_population_of_molecules_found
Integrity (uniformity) of mRNA tested using real-time quantitative polymerase chain reaction (rtqPCR):
not_tested
Integrity (uniformity) of mRNA tested using RNAi:
not_tested
Integrity (uniformity) of mRNA tested using S1 nuclease mapping:
not_tested
Cryptic promoter presence was confirmed by expression from a promoter-less plasmid:
no_promoter_confirmed
Cryptic promoter presence was confirmed in an experimental setup involving inducible promoter:
not_tested
Integrity (uniformity) of mRNA molecules or possible promoter presence expressed in vivo was tested using another method, please specify in Remarks:
not_tested
The organism used:
Homo sapiens HepG2 (ATCC HB-8065)
The abbreviated name of the donor gene or virus from which this IRES was excised and inserted into the plasmid:
c-IAP1
The origin of IRES in the plasmid:
  nuclear
The donor organism of the IRES segment:
Homo sapiens HeLa (ATCC CCL-2)
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:
pRL-cIAP1-FL.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):
  194-1129
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):
  2279-3931
Remarks:
The intercistronic region contains incomplete 5'-UTR of c-IAP1 (-1115 to -1) followed by some 19 b as a
remnant of HCV ORF after mutation.
Citations:
Van Eden M. E., Byrd M. P., Sherrill K. W., Lloyd R. E. (2004) Translation of cellular inhibitor of apoptosis protein 1 (c-IAP1) mRNA is IRES mediated and regulated during cell stress. RNA. 10(3):469-481
IRESs:
IRES:
Version: 1 Last change: 2008-10-23 16:22:52
Originaly submitted by: Martin Mokrejš Reviewed by: Martin Mokrejš
The IRES name:
  c-IAP1_-1178/-64
The functional status of IRES:
  functional
The IRES absolute position (the range includes START and STOP codons or their equivalents):
  1145-2259
How IRES boundaries were determined:
experimentally_determined
5'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  16
3'-end of IRES relative to last base of the STOP codon of the upstream ORF:
  1130
5'-end of IRES relative to first base of the START codon of the downstream ORF:
  -1134
3'-end of IRES relative to first base of the START codon of the downstream ORF:
  -20
The sequence of IRES region aligned to its secondary structure (if available):


Remarks:
The results are biased by a minor splice variant.
Citations:
Van Eden M. E., Byrd M. P., Sherrill K. W., Lloyd R. E. (2004) Translation of cellular inhibitor of apoptosis protein 1 (c-IAP1) mRNA is IRES mediated and regulated during cell stress. RNA. 10(3):469-481
The translation experiments:
Translation results:
IRESite Translation Id: 331
Version: 1 Last change: 2007-02-16 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:
Homo sapiens HepG2 (ATCC HB-8065)
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  1117.600
Name of IRES used as the positive control:
  HCV_type_1b
Name of the plasmid used as the positive control.
pRL-HCV-FL
Name of the plasmid used as the negative control.
p-RL-FL
IRESite Id of the plasmid used as positive control.
  131
IRESite Id of the plasmid used as negative control.
  119
The relative translation efficiency in % of the positive control:
  100.000
The relative translation efficiency in % of the negative control:
  52.900
The size (length) of intercistronic region in the positive control:
419
The size (length) of intercistronic region in the negative control:
28
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:
Fig. 4b
Citations:
Van Eden M. E., Byrd M. P., Sherrill K. W., Lloyd R. E. (2004) Translation of cellular inhibitor of apoptosis protein 1 (c-IAP1) mRNA is IRES mediated and regulated during cell stress. RNA. 10(3):469-481
Translation results:
IRESite Translation Id: 332
Version: 1 Last change: 2007-02-16 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:
Homo sapiens HeLa S3 (ATCC CCL-2.2)
The temperature (in degrees of Celsia):
37
The relative translation efficiency in % of this IRES:
  300.000
Name of IRES used as the positive control:
  HCV_type_1b
Name of the plasmid used as the positive control.
pRL-HCV-FL
Name of the plasmid used as the negative control.
p-RL-FL
IRESite Id of the plasmid used as positive control.
  131
IRESite Id of the plasmid used as negative control.
  119
The relative translation efficiency in % of the positive control:
  100.000
The relative translation efficiency in % of the negative control:
  50.000
The size (length) of intercistronic region in the positive control:
419
The size (length) of intercistronic region in the negative control:
28
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:
Fig. 4b
Citations:
Van Eden M. E., Byrd M. P., Sherrill K. W., Lloyd R. E. (2004) Translation of cellular inhibitor of apoptosis protein 1 (c-IAP1) mRNA is IRES mediated and regulated during cell stress. RNA. 10(3):469-481
Last change to the database: 2015-04-16 16:45:23 GMT+1