IRESite_Id:211 characteristics of CVB3 IRES studied in pRSTF-CVB3 plasmid

IRESite: The database of experimentally verified IRES structures

The nucleic acid data:

IRESite Id: `211`	Version: `8`
Originaly submitted by: Martin Mokrejš
Reviewed by: Martin Mokrejš	Last change: `2008-02-01 00:00:00`

IRESite record type:

plasmid_with_promoter_and_putative_IRES_without_translational_characterization

The shape of the nucleic acid molecule translated:
linear The quality of the mRNA/+RNA sequence:
hopefully_full-length_mRNA

The mRNA/+RNA description:

Putative in vivo transcript from SV40 promoter with the chimeric intron spliced out and containing
renilla_luciferase - hairpin_from_multiple_cloning_site - CVB3_nt1-750 - 45_nt_of_FMDV_2A_protease -
firefly_luciferase - late_SV40_polyA_signal. A fusion protein CVB3+FMDV+Fluc should be translated from the
second cistron (N-terminal 31 additional aminoacids fused to FLuc).

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

attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agcttgattc ttctgacaca acagtctcga acttaagctg cagaagttgg 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

tcgtgaggca ctgggcaggt gtccactccc agttcaatta cagctcttaa ggctagagta cTTAATACGA CTCACTATAG Gctagccacc atGacttcga 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

aagtttatga tccagaacaa aggaaacgga tgataactgg tccgcagtgg tgggccagat gtaaacaaat gaatgttctt gattcattta ttaattatta 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

tgattcagaa aaacatgcag aaaatgctgt tattttttta catggtaacg cggcctcttc ttatttatgg cgacatgttg tgccacatat tgagccagta 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

gcgcggtgta ttataccaga ccttattggt atgggcaaat caggcaaatc tggtaatggt tcttataggt tacttgatca ttacaaatat cttactgcat 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

ggtttgaact tcttaattta ccaaagaaga tcatttttgt cggccatgat tggggtgctt gtttggcatt tcattatagc tatgagcatc aagataagat 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

caaagcaata gttcacgctg aaagtgtagt agatgtgatt gaatcatggg atgaatggcc tgatattgaa gaagatattg cgttgatcaa atctgaagaa 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

ggagaaaaaa tggttttgga gaataacttc ttcgtggaaa ccatgttgcc atcaaaaatc atgagaaagt tagaaccaga agaatttgca gcatatcttg 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

aaccattcaa agagaaaggt gaagttcgtc gtccaacatt atcatggcct cgtgaaatcc cgttagtaaa aggtggtaaa cctgacgttg tacaaattgt 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

taggaattat aatgcttatc tacgtgcaag tgatgattta ccaaaaatgt ttattgaatc ggacccagga ttcttttcca atgctattgt tgaaggtgcc 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

aagaagtttc ctaatactga atttgtcaaa gtaaaaggtc ttcatttttc gcaagaagat gcacctgatg aaatgggaaa atatatcaaa tcgttcgttg 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

agcgagttct caaaaatgaa caaatGTCGA cctaggcttt tgcaaaAAGC TTctcgaggc gaggtggcga ccgcgcatgc gcggtcgcca cctcgcctcg 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

agAAGCTTAA ttaaaacagc ctgtgggttg atcccaccca cagggcccat tgggcgctag cactctggta tcacggtacc tttgtgcgcc tgttttatac 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

cccctccccc aactgtaact tagaagtaac acacaccgat caacagtcag cgtggcacac cagccacgtt ttgatcaagc acttctgtta ccccggactg 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

agtatcaata gactgctcac gcggttgaag gagaaagcgt tcgttatccg gccaactact tcgaaaaacc tagtaacacc gtggaagttg cagagtgttt 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

cgctcagcac taccccagtg tagatcaggt cgatgagtca ccgcattccc cacgggcgac cgtggcggtg gctgcgttgg cggcctgccc atggggaaac 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

ccatgggacg ctctaataca gacatggtgc gaagagtcta ttgagctagt tggtagtcct ccggcccctg aatgcggcta atcctaactg cggagcacac 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

accctcaagc cagagggcag tgtgtcgtaa cgggcaactc tgcagcggaa ccgactactt tgggtgtccg tgtttcattt tattcctata ctggctgctt 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

atggtgacaa ttgagagatt gttaccatat agctattgga ttggccatcc ggtgaccaat agagctatta tatatctctt tgttgggttt ataccactta 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

gcttgaaaga ggttaaaaca ttacaattca ttgttaagtt gaatacagca aaatgggaGC GGCCGCaaat tttgaccttc tcaagttggc gggagacgtc 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

gagtccaacc ctGGGCCCct aggagatcta gcgctGgatc GATCCATGGa agacgccaaa aacataaaga aaggcccggc gccattctat ccgctggaag 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

atggaaccgc tggagagcaa ctgcataagg ctatgaagag atacgccctg gttcctggaa caattgcttt tacagatgca catatcgagg tggacatcac 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

ttacgctgag tacttcgaaa tgtccgttcg gttggcagaa gctatgaaac gatatgggct gaatacaaat cacagaatcg tcgtatgcag tgaaaactct 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

cttcaattct ttatgccggt gttgggcgcg ttatttatcg gagttgcagt tgcgcccgcg aacgacattt ataatgaacg tgaattgctc aacagtatgg 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

gcatttcgca gcctaccgtg gtgttcgttt ccaaaaaggg gttgcaaaaa attttgaacg tgcaaaaaaa gctcccaatc atccaaaaaa ttattatcat 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

ggattctaaa acggattacc agggatttca gtcgatgtac acgttcgtca catctcatct acctcccggt tttaatgaat acgattttgt gccagagtcc 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

ttcgataggg acaagacaat tgcactgatc atgaactcct ctggatctac tggtctgcct aaaggtgtcg ctctgcctca tagaactgcc tgcgtgagat 
^2601      ^2611      ^2621      ^2631      ^2641      ^2651      ^2661      ^2671      ^2681      ^2691      

tctcgcatgc cagagatcct atttttggca atcaaatcat tccggatact gcgattttaa gtgttgttcc attccatcac ggttttggaa tgtttactac 
^2701      ^2711      ^2721      ^2731      ^2741      ^2751      ^2761      ^2771      ^2781      ^2791      

actcggatat ttgatatgtg gatttcgagt cgtcttaatg tatagatttg aagaagagct gtttctgagg agccttcagg attacaagat tcaaagtgcg 
^2801      ^2811      ^2821      ^2831      ^2841      ^2851      ^2861      ^2871      ^2881      ^2891      

ctgctggtgc caaccctatt ctccttcttc gccaaaagca ctctgattga caaatacgat ttatctaatt tacacgaaat tgcttctggt ggcgctcccc 
^2901      ^2911      ^2921      ^2931      ^2941      ^2951      ^2961      ^2971      ^2981      ^2991      

tctctaagga agtcggggaa gcggttgcca agaggttcca tctgccaggt atcaggcaag gatatgggct cactgagact acatcagcta ttctgattac 
^3001      ^3011      ^3021      ^3031      ^3041      ^3051      ^3061      ^3071      ^3081      ^3091      

acccgagggg gatgataaac cgggcgcggt cggtaaagtt gttccatttt ttgaagcgaa ggttgtggat ctggataccg ggaaaacgct gggcgttaat 
^3101      ^3111      ^3121      ^3131      ^3141      ^3151      ^3161      ^3171      ^3181      ^3191      

caaagaggcg aactgtgtgt gagaggtcct atgattatgt ccggttatgt aaacaatccg gaagcgacca acgccttgat tgacaaggat ggatggctac 
^3201      ^3211      ^3221      ^3231      ^3241      ^3251      ^3261      ^3271      ^3281      ^3291      

attctggaga catagcttac tgggacgaag acgaacactt cttcatcgtt gaccgcctga agtctctgat taagtacaaa ggctatcagg tggctcccgc 
^3301      ^3311      ^3321      ^3331      ^3341      ^3351      ^3361      ^3371      ^3381      ^3391      

tgaattggaa tccatcttgc tccaacaccc caacatcttc gacgcaggtg tcgcaggtct tcccgacgat gacgccggtg aacttcccgc cgccgttgtt 
^3401      ^3411      ^3421      ^3431      ^3441      ^3451      ^3461      ^3471      ^3481      ^3491      

gttttggagc acggaaagac gatgacggaa aaagagatcg tggattacgt cgccagtcaa gtaacaaccg cgaaaaagtt gcgcggagga gttgtgtttg 
^3501      ^3511      ^3521      ^3531      ^3541      ^3551      ^3561      ^3571      ^3581      ^3591      

tggacgaagt accgaaaggt cttaccggaa aactcgacgc aagaaaaatc agagagatcc tcataaaggc caagaagggc ggaaagatcg ccgtgtaatT 
^3601      ^3611      ^3621      ^3631      ^3641      ^3651      ^3661      ^3671      ^3681      ^3691      

CTAGAgcggc cggccgcttc gagcagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa 
^3701      ^3711      ^3721      ^3731      ^3741      ^3751      ^3761      ^3771      ^3781      ^3791      

atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg 
^3801      ^3811      ^3821      ^3831      ^3841      ^3851      ^3861      ^3871      ^3881      ^3891      

tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggta
^3901      ^3911      ^3921      ^3931      ^3941

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_sequence_and_should_match_experiment_maybe_except_both_UTRs

The name of the plasmid:
pRSTF-CVB3 The name of the promoter used to express this mRNA:
SV40

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 (not convincing)

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:
unclear_result

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:
not_tested

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 HeLa (ATCC CCL-2)