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-fusion reporter genes separated by GBV-B IRES segment with 15 nts mutated in a way to
disrupt potential stem loop (incl. ATG).
The mRNA/+RNA sequence represented in the +DNA notation:
Credibility of mRNA sequence: end-to-end_sequence_reverse_engineered_and_should_match_experiment
The description of the protein encoded in this ORF: Chloramphenicol acetyltransferase with additional N-terminal 5 aminoacid residues (mutated nucleotides to
disrupt secondary structure stem loop).
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): 1468-2151
Comparing Fig. 7b and 7c shows that the "randomly" created sequence could have a promoter activity in vivo.
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.
The relative translation efficiency in % of this IRES: 142.000
Name of IRES used as the positive control: GBV-B+14
Name of the plasmid used as the positive control. pBL-RLuc-GBB+15-CATT
IRESite Id of the plasmid used as positive control. 178
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
The mutated GBV-B IRES with modified trailing 15nts seems to be active only in vivo. It is highly possible the
sequence introduced to disrupt secondary structure stem loop has a promoter activity (Fig. 7c).