IRESite_Id:614 characteristics of (GAAA)16 IRES studied in H-GFP-(GAAA)16-GUS plasmid

IRESite: The database of experimentally verified IRES structures

The nucleic acid data:

IRESite Id: `614`	Version: `0`
Originaly submitted by: Martin Mokrejš	Submission date: `2009-08-26 01:24:36`
Reviewed by: Martin Mokrejš

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:

T7-promoter derived transcript containing synthetic IRES consisting of GAAA motif repeated 16 times inserted
between GFP and GUS reporters.

The mRNA/+RNA sequence represented in the +DNA notation:

ATGGTGAGCA AGGGCGAGGA GCTGTTCACC GGGGTGGTGC CCATCCTGGT CGAGCTGGAC GGCGACGTAA ACGGCCACAA GTTCAGCGTG TCCGGCGAGG 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

GCGAGGGCGA TGCCACCTAC GGCAAGCTGA CCCTGAAGTT CATCTGCACC ACCGGCAAGC TGCCCGTGCC CTGGCCCACC CTCGTGACCA CCTTCACCTA 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

CGGCGTGCAG TGCTTCAGCC GCTACCCCGA CCACATGAAG CAGCACGACT TCTTCAAGTC CGCCATGCCC GAAGGCTACG TCCAGGAGCG CACCATCTTC 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

TTCAAGGACG ACGGCAACTA CAAGACCCGC GCCGAGGTGA AGTTCGAGGG CGACACCCTG GTGAACCGCA TCGAGCTGAA GGGCATCGAC TTCAAGGAGG 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

ACGGCAACAT CCTGGGGCAC AAGCTGGAGT ACAACTACAA CAGCCACAAC GTCTATATCA TGGCCGACAA GCAGAAGAAC GGCATCAAGG TGAACTTCAA 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

GATCCGCCAC AACATCGAGG ACGGCAGCGT GCAGCTCGCC GACCACTACC AGCAGAACAC CCCCATCGGC GACGGCCCCG TGCTGCTGCC CGACAACCAC 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

TACCTGAGCA CCCAGTCCGC CCTGAGCAAA GACCCCAACG AGAAGCGCGA TCACATGGTC CTGCTGGAGT TCGTGACCGC CGCCGGGATC ACTCACGGCA 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

TGGACGAGCT GTACAAGTAA GCTTAGAAAG AAAGAAAGAA AGAAAGAAAG AAACTGCAGC CGCGCCTGCA GAAAGAAAGA AAGAAAGAAA GAAAGAAAGA 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

AAGAATTCAT GGTCCGTCCT GTAGAAACCC CAACCCGTGA AATCAAAAAA CTCGACGGCC TGTGGGCATT CAGTCTGGAT CGCGAAAACT GTGGAATTGA 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

TCAGCGTTGG TGGGAAAGCG CGTTACAAGA AAGCCGGGCA ATTGCTGTGC CAGGCAGTTT TAACGATCAG TTCGCCGATG CAGATATTCG TAATTATGCG 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

GGCAACGTCT GGTATCAGCG CGAAGTCTTT ATACCGAAAG GTTGGGCAGG CCAGCGTATC GTGCTGCGTT TCGATGCGGT CACTCATTAC GGCAAAGTGT 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

GGGTCAATAA TCAGGAAGTG ATGGAGCATC AGGGCGGCTA TACGCCATTT GAAGCCGATG TCACGCCGTA TGTTATTGCC GGGAAAAGTG TACGTATCAC 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

CGTTTGTGTG AACAACGAAC TGAACTGGCA GACTATCCCG CCGGGAATGG TGATTACCGA CGAAAACGGC AAGAAAAAGC AGTCTTACTT CCATGATTTC 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

TTTAACTATG CCGGAATCCA TCGCAGCGTA ATGCTCTACA CCACGCCGAA CACCTGGGTG GACGATATCA CCGTGGTGAC GCATGTCGCG CAAGACTGTA 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

ACCACGCGTC TGTTGACTGG CAGGTGGTGG CCAATGGTGA TGTCAGCGTT GAACTGCGTG ATGCGGATCA ACAGGTGGTT GCAACTGGAC AAGGCACTAG 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

CGGGACTTTG CAAGTGGTGA ATCCGCACCT CTGGCAACCG GGTGAAGGTT ATCTCTATGA ACTGTGCGTC ACAGCCAAAA GCCAGACAGA GTGTGATATC 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

TACCCGCTTC GCGTCGGCAT CCGGTCAGTG GCAGTGAAGG GCGAACAGTT CCTGATTAAC CACAAACCGT TCTACTTTAC TGGCTTTGGT CGTCATGAAG 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

ATGCGGACTT ACGTGGCAAA GGATTCGATA ACGTGCTGAT GGTGCACGAC CACGCATTAA TGGACTGGAT TGGGGCCAAC TCCTACCGTA CCTCGCATTA 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

CCCTTACGCT GAAGAGATGC TCGACTGGGC AGATGAACAT GGCATCGTGG TGATTGATGA AACTGCTGCT GTCGGCTTTA ACCTCTCTTT AGGCATTGGT 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

TTCGAAGCGG GCAACAAGCC GAAAGAACTG TACAGCGAAG AGGCAGTCAA CGGGGAAACT CAGCAAGCGC ACTTACAGGC GATTAAAGAG CTGATAGCGC 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

GTGACAAAAA CCACCCAAGC GTGGTGATGT GGAGTATTGC CAACGAACCG GATACCCGTC CGCAAGGTGC ACGGGAATAT TTCGCGCCAC TGGCGGAAGC 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

AACGCGTAAA CTCGACCCGA CGCGTCCGAT CACCTGCGTC AATGTAATGT TCTGCGACGC TCACACCGAT ACCATCAGCG ATCTCTTTGA TGTGCTGTGC 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

CTGAACCGTT ATTACGGATG GTATGTCCAA AGCGGCGATT TGGAAACGGC AGAGAAGGTA CTGGAAAAAG AACTTCTGGC CTGGCAGGAG AAACTGCATC 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

AGCCGATTAT CATCACCGAA TACGGCGTGG ATACGTTAGC CGGGCTGCAC TCAATGTACA CCGACATGTG GAGTGAAGAG TATCAGTGTG CATGGCTGGA 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

TATGTATCAC CGCGTCTTTG ATCGCGTCAG CGCCGTCGTC GGTGAACAGG TATGGAATTT CGCCGATTTT GCGACCTCGC AAGGCATATT GCGCGTTGGC 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

GGTAACAAGA AAGGGATCTT CACTCGCGAC CGCAAACCGA AGTCGGCGGC TTTTCTGCTG CAAAAACGCT GGACTGGCAT GAACTTCGGT GAAAAACCGC 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

AGCAGGGAGG CAAACAATGA
^2601      ^2611

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:

end-to-end_sequence_reverse_engineered_and_should_match_experiment

The name of the plasmid:
H-GFP-(GAAA)16-GUS 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 origin of IRES in the plasmid:

engineered

The DNA sequence of the plasmid in (+) orientation annotated by its secondary structure:

ATGGTGAGCA AGGGCGAGGA GCTGTTCACC GGGGTGGTGC CCATCCTGGT CGAGCTGGAC GGCGACGTAA ACGGCCACAA GTTCAGCGTG TCCGGCGAGG 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

GCGAGGGCGA TGCCACCTAC GGCAAGCTGA CCCTGAAGTT CATCTGCACC ACCGGCAAGC TGCCCGTGCC CTGGCCCACC CTCGTGACCA CCTTCACCTA 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

CGGCGTGCAG TGCTTCAGCC GCTACCCCGA CCACATGAAG CAGCACGACT TCTTCAAGTC CGCCATGCCC GAAGGCTACG TCCAGGAGCG CACCATCTTC 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

TTCAAGGACG ACGGCAACTA CAAGACCCGC GCCGAGGTGA AGTTCGAGGG CGACACCCTG GTGAACCGCA TCGAGCTGAA GGGCATCGAC TTCAAGGAGG 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

ACGGCAACAT CCTGGGGCAC AAGCTGGAGT ACAACTACAA CAGCCACAAC GTCTATATCA TGGCCGACAA GCAGAAGAAC GGCATCAAGG TGAACTTCAA 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

GATCCGCCAC AACATCGAGG ACGGCAGCGT GCAGCTCGCC GACCACTACC AGCAGAACAC CCCCATCGGC GACGGCCCCG TGCTGCTGCC CGACAACCAC 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

TACCTGAGCA CCCAGTCCGC CCTGAGCAAA GACCCCAACG AGAAGCGCGA TCACATGGTC CTGCTGGAGT TCGTGACCGC CGCCGGGATC ACTCACGGCA 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

TGGACGAGCT GTACAAGTAA GCTTAGAAAG AAAGAAAGAA AGAAAGAAAG AAACTGCAGC CGCGCCTGCA GAAAGAAAGA AAGAAAGAAA GAAAGAAAGA 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

AAGAATTCAT GGTCCGTCCT GTAGAAACCC CAACCCGTGA AATCAAAAAA CTCGACGGCC TGTGGGCATT CAGTCTGGAT CGCGAAAACT GTGGAATTGA 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

TCAGCGTTGG TGGGAAAGCG CGTTACAAGA AAGCCGGGCA ATTGCTGTGC CAGGCAGTTT TAACGATCAG TTCGCCGATG CAGATATTCG TAATTATGCG 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

GGCAACGTCT GGTATCAGCG CGAAGTCTTT ATACCGAAAG GTTGGGCAGG CCAGCGTATC GTGCTGCGTT TCGATGCGGT CACTCATTAC GGCAAAGTGT 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

GGGTCAATAA TCAGGAAGTG ATGGAGCATC AGGGCGGCTA TACGCCATTT GAAGCCGATG TCACGCCGTA TGTTATTGCC GGGAAAAGTG TACGTATCAC 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

CGTTTGTGTG AACAACGAAC TGAACTGGCA GACTATCCCG CCGGGAATGG TGATTACCGA CGAAAACGGC AAGAAAAAGC AGTCTTACTT CCATGATTTC 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

TTTAACTATG CCGGAATCCA TCGCAGCGTA ATGCTCTACA CCACGCCGAA CACCTGGGTG GACGATATCA CCGTGGTGAC GCATGTCGCG CAAGACTGTA 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

ACCACGCGTC TGTTGACTGG CAGGTGGTGG CCAATGGTGA TGTCAGCGTT GAACTGCGTG ATGCGGATCA ACAGGTGGTT GCAACTGGAC AAGGCACTAG 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

CGGGACTTTG CAAGTGGTGA ATCCGCACCT CTGGCAACCG GGTGAAGGTT ATCTCTATGA ACTGTGCGTC ACAGCCAAAA GCCAGACAGA GTGTGATATC 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

TACCCGCTTC GCGTCGGCAT CCGGTCAGTG GCAGTGAAGG GCGAACAGTT CCTGATTAAC CACAAACCGT TCTACTTTAC TGGCTTTGGT CGTCATGAAG 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

ATGCGGACTT ACGTGGCAAA GGATTCGATA ACGTGCTGAT GGTGCACGAC CACGCATTAA TGGACTGGAT TGGGGCCAAC TCCTACCGTA CCTCGCATTA 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

CCCTTACGCT GAAGAGATGC TCGACTGGGC AGATGAACAT GGCATCGTGG TGATTGATGA AACTGCTGCT GTCGGCTTTA ACCTCTCTTT AGGCATTGGT 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

TTCGAAGCGG GCAACAAGCC GAAAGAACTG TACAGCGAAG AGGCAGTCAA CGGGGAAACT CAGCAAGCGC ACTTACAGGC GATTAAAGAG CTGATAGCGC 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

GTGACAAAAA CCACCCAAGC GTGGTGATGT GGAGTATTGC CAACGAACCG GATACCCGTC CGCAAGGTGC ACGGGAATAT TTCGCGCCAC TGGCGGAAGC 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

AACGCGTAAA CTCGACCCGA CGCGTCCGAT CACCTGCGTC AATGTAATGT TCTGCGACGC TCACACCGAT ACCATCAGCG ATCTCTTTGA TGTGCTGTGC 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

CTGAACCGTT ATTACGGATG GTATGTCCAA AGCGGCGATT TGGAAACGGC AGAGAAGGTA CTGGAAAAAG AACTTCTGGC CTGGCAGGAG AAACTGCATC 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

AGCCGATTAT CATCACCGAA TACGGCGTGG ATACGTTAGC CGGGCTGCAC TCAATGTACA CCGACATGTG GAGTGAAGAG TATCAGTGTG CATGGCTGGA 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

TATGTATCAC CGCGTCTTTG ATCGCGTCAG CGCCGTCGTC GGTGAACAGG TATGGAATTT CGCCGATTTT GCGACCTCGC AAGGCATATT GCGCGTTGGC 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

GGTAACAAGA AAGGGATCTT CACTCGCGAC CGCAAACCGA AGTCGGCGGC TTTTCTGCTG CAAAAACGCT GGACTGGCAT GAACTTCGGT GAAAAACCGC 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

AGCAGGGAGG CAAACAATGA
^2601      ^2611

GenBank formatted file with annotated plasmid sequence hyperlinked from vector image map:

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:
GFP

The description of the protein encoded in this ORF:
green fluorescent 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):

1-720

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):

809-2620

Citations:

Dorokhov YL, Skulachev MV, Ivanov PA, Zvereva SD, Tjulkina LG, Merits A, Gleba YY, Hohn T, Atabekov JG (2002) Polypurine (A)-rich sequences promote cross-kingdom conservation of internal ribosome entry. Proc. Natl. Acad. Sci. U.S.A. 8(99):5301-5306

IRESs:

IRES:

Version: `0`
Originaly submitted by: Martin Mokrejš	Reviewed by: Martin Mokrejš
The IRES name: `(GAAA)16`	The functional status of IRES: `functional`

The IRES absolute position (the range includes START and STOP codons or their equivalents):

726-805

How IRES boundaries were determined:
experimentally_determined

5'-end of IRES relative to last base of the STOP codon of the upstream ORF: `6`	3'-end of IRES relative to last base of the STOP codon of the upstream ORF: `85`
5'-end of IRES relative to first base of the START codon of the downstream ORF: `-83`	3'-end of IRES relative to first base of the START codon of the downstream ORF: `-4`

The sequence of IRES region aligned to its secondary structure (if available):

GAAAGAAAGA AAGAAAGAAA GAAAGAAACT GCAGCCGCGC CTGCAGAAAG AAAGAAAGAA AGAAAGAAAG AAAGAAAGAA
^726       ^736       ^746       ^756       ^766       ^776       ^786       ^796

Citations:

The translation experiments:

Translation results:

IRESite Translation Id: 642

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):
25

The relative translation efficiency in % of this IRES:

120.000

Name of IRES used as the positive control:

crTMV_IREScp

Name of the plasmid used as the positive control. `H-GFP-IREScp-GUS`
`IRESite Id` of the plasmid used as positive control. `609`
The relative translation efficiency in % of the positive control: `100.000`
The size (length) of intercistronic region in the positive control: `177`

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 from Figure 4A.

Citations:

Translation results:

IRESite Translation Id: 643

Version: `0`
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 (ATCC CCL-2)

The temperature (in degrees of Celsia):
37

The relative translation efficiency in % of this IRES:

140.000

Name of IRES used as the positive control:

EMCV-R

Name of the plasmid used as the positive control. `H-GFP-IRESemcv-GUS`
`IRESite Id` of the plasmid used as positive control. `603`
The relative translation efficiency in % of the positive control: `100.000`
The size (length) of intercistronic region in the positive control: `595`

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:

Data from Figure 4C. The cells were infected with recombinant vaccinia virus.

Citations:

Last change to the database: 2019-03-18 09:32:49 GMT+1