IRESite_Id:559 characteristics of eIF4G_-368_-12 IRES studied in pGLuc-eIF4G-GFP plasmid

IRESite: The database of experimentally verified IRES structures

The nucleic acid data:

IRESite Id: `559`	Version: `4`
Originaly submitted by: Martin Mokrejš	Submission date: `2009-03-13 17:27:33`
Reviewed by: Martin Mokrejš	Last change: `2009-09-02 22:13:19`

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:
our_best_guess

The mRNA/+RNA description:

Putative in vivo transcript driven from CMV IE promoter and terminated after the SV40 late poly(A) signal
containing GLuc (Gaussia luciferase) and GFP cistrons separated by the putative eIF4G IRES.

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

GTCAGATCAC TAGAAGCTTA TCAGACGACA TCACTCGAGA CTAACTCACC GGAATTCCAA GCTGGCAACT TCCGGAAGCT GCAAGCCACC ATGGGAGTCA 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

AAGTCCTGTT CGCTCTGATC TGCATCGCTG TGGCCGAGGC CAAGCCCACC GAGAACAACG AAGACTTCAA CATCGTCGCC GTGGCCAGCA ACTTCGCAAC 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

CACTGATCTG GACGCTGACA GAGGCAAACT GCCTGGCAAG AAGCTGCCTC TCGAAGTCCT GAAAGAGATG GAAGCCAACG CTCGGAAGGC TGGCTGCACA 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

AGAGGCTGCC TGATCTGCCT GTCCCACATC AAATGCACAC CCAAGATGAA GAAATTCATC CCTGGACGCT GCCACACCTA CGAAGGCGAC AAAGAATCCG 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

CACAGGGAGG CATTGGAGAA GCCATTGTCG ACATTCCCGA GATTCCTGGA TTCAAGGACC TGGAGCCTAT GGAGCAGTTC ATCGCACAAG TCGACCTCTG 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

CGTGGACTGC ACAACTGGAT GCCTCAAAGG CCTCGCTAAT GTCCAATGCT CTGACCTGCT CAAGAAATGG CTCCCACAGC GCTGCGCTAC CTTCGCAAGC 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

AAGATCCAGG GCCAAGTCGA TAAGATCAAG GGAGCAGGCG GAGACTAAAC CTAGTTTAAA CGCGTCTAGA TGGGGGTCCT GGGCCCCAGG gtgtgcagcc 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

actgacttgg ggactgctgg tggggtaggg atgagggagg gaggggcatt gtgatgtaca gggctgctct gtgagatcaa gggtctctta agggtgggag 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

ctggggcagg gactacgaga gcagccagat gggctgaaag tggaactcaa ggggtttctg gcacctacct acctgcttcc cgctgggggg tggggagttg 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

gcccagagtc ttaagattgg ggcagggtgg agaggtgggc tcttcctgct tcccactcat cttatagctt tctttcccca gATCCGAATT CGAGATCCAA 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

ACCAAGGAGG AAAGGATATC ACGTACGATG GTGAGCAAGG GCGAGGAGCT GTTCACCGGG GTGGTGCCCA TCCTGGTCGA GCTGGACGGC GACGTAAACG 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

GCCACAAGTT CAGCGTGTCC GGCGAGGGCG AGGGCGATGC CACCTACGGC AAGCTGACCC TGAAGTTCAT CTGCACCACC GGCAAGCTGC CCGTGCCCTG 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

GCCCACCCTC GTGACCACCT TGACCTACGG CGTGCAGTGC TTCGCCCGCT ACCCCGACCA CATGAAGCAG CACGACTTCT TCAAGTCCGC CATGCCCGAA 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

GGCTACGTCC AGGAGCGCAC CATCTTCTTC AAGGACGACG GCAACTACAA GACCCGCGCC GAGGTGAAGT TCGAGGGCGA CACCCTGGTG AACCGCATCG 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

AGCTGAAGGG CATCGACTTC AAGGAGGACG GCAACATCCT GGGGCACAAG CTGGAGTACA ACTACAACAG CCACAAGGTC TATATCACCG CCGACAAGCA 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

GAAGAACGGC ATCAAGGTGA ACTTCAAGAC CCGCCACAAC ATCGAGGACG GCAGCGTGCA GCTCGCCGAC CACTACCAGC AGAACACCCC CATCGGCGAC 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

GGCCCCGTGC TGCTGCCCGA CAACCACTAC CTGAGCACCC AGTCCGCCCT GAGCAAAGAC CCCAACGAGA AGCGCGATCA CATGGTCCTG CTGGAGTTCG 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

TGACCGCCGC CGGGATCACT CTCGGCATGG ACGAGCTGTA CAAGTAGCGG CCCAATCTAG AGTCGGGGCG GCCGGCCGCT TCGAGCAGAC ATGATAAGAT 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

ACATTGATGA GTTTGGACAA ACCACAACTA GAATGCAGTG AAAAAAATGC TTTATTTGTG AAATTTGTGA TGCTATTGCT TTATTTGTAA CCATTATAAG 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

CTGCAATAAA CAAGTTAACA ACAACAATTG CATTCATTTT ATGTTTCAGG TTCAGGGGGA GGTGTGGGAG GTTTTTTAAA GCAAGTAAAA CCTCTACAAA 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

TGTGGTA
^2001

Credibility of mRNA sequence:

end-to-end_sequence_reverse_engineered_and_should_match_experiment

The name of the plasmid:
pGLuc-eIF4G-GFP The name of the promoter used to express this mRNA:
CMV_IE

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:

not tested

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:
not_tested

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)

The abbreviated name of the donor gene or virus from which this IRES was excised and inserted into the plasmid:
eIF4G

The origin of IRES in the plasmid:

nuclear

The donor organism of the IRES segment:

Homo sapiens

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

TCAATATTGG CCATTAGCCA TATTATTCAT TGGTTATATA GCATAAATCA ATATTGGCTA TTGGCCATTG CATACGTTGT ATCTATATCA TAATATGTAC 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

ATTTATATTG GCTCATGTCC AATATGACCG CCATGTTGGC ATTGATTATT GACTAGTTAT TAATAGTAAT CAATTACGGG GTCATTAGTT CATAGCCCAT 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

ATATGGAGTT CCGCGTTACA TAACTTACGG TAAATGGCCC GCCTGGCTGA CCGCCCAACG ACCCCCGCCC ATTGACGTCA ATAATGACGT ATGTTCCCAT 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

AGTAACGCCA ATAGGGACTT TCCATTGACG TCAATGGGTG GAGTATTTAC GGTAAACTGC CCACTTGGCA GTACATCAAG TGTATCATAT GCCAAGTCCG 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

CCCCCTATTG ACGTCAATGA CGGTAAATGG CCCGCCTGGC ATTATGCCCA GTACATGACC TTACGGGACT TTCCTACTTG GCAGTACATC TACGTATTAG 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

TCATCGCTAT TACCATGGTG ATGCGGTTTT GGCAGTACAC CAATGGGCGT GGATAGCGGT TTGACTCACG GGGATTTCCA AGTCTCCACC CCATTGACGT 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

CAATGGGAGT TTGTTTTGGC ACCAAAATCA ACGGGACTTT CCAAAATGTC GTAATAACCC CGCCCCGTTG ACGCAAATGG GCGGTAGGCG TGTACGGTGG 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

GAGGTCTATA TAAGCAGAGC TCGTTTAGTG AACCGTCAGA TCACTAGAAG CTTATCAGAC GACATCACTC GAGACTAACT CACCGGAATT CCAAGCTGGC 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

AACTTCCGGA AGCTGCAAGC CACCATGGGA GTCAAAGTCC TGTTCGCTCT GATCTGCATC GCTGTGGCCG AGGCCAAGCC CACCGAGAAC AACGAAGACT 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

TCAACATCGT CGCCGTGGCC AGCAACTTCG CAACCACTGA TCTGGACGCT GACAGAGGCA AACTGCCTGG CAAGAAGCTG CCTCTCGAAG TCCTGAAAGA 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

GATGGAAGCC AACGCTCGGA AGGCTGGCTG CACAAGAGGC TGCCTGATCT GCCTGTCCCA CATCAAATGC ACACCCAAGA TGAAGAAATT CATCCCTGGA 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

CGCTGCCACA CCTACGAAGG CGACAAAGAA TCCGCACAGG GAGGCATTGG AGAAGCCATT GTCGACATTC CCGAGATTCC TGGATTCAAG GACCTGGAGC 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

CTATGGAGCA GTTCATCGCA CAAGTCGACC TCTGCGTGGA CTGCACAACT GGATGCCTCA AAGGCCTCGC TAATGTCCAA TGCTCTGACC TGCTCAAGAA 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

ATGGCTCCCA CAGCGCTGCG CTACCTTCGC AAGCAAGATC CAGGGCCAAG TCGATAAGAT CAAGGGAGCA GGCGGAGACT AAACCTAGTT TAAACGCGTC 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

TAGATGGGGG TCCTGGGCCC CAGGgtgtgc agccactgac ttggggactg ctggtggggt agggatgagg gagggagggg cattgtgatg tacagggctg 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

ctctgtgaga tcaagggtct cttaagggtg ggagctgggg cagggactac gagagcagcc agatgggctg aaagtggaac tcaaggggtt tctggcacct 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

acctacctgc ttcccgctgg ggggtgggga gttggcccag agtcttaaga ttggggcagg gtggagaggt gggctcttcc tgcttcccac tcatcttata 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

gctttctttc cccagATCCG AATTCGAGAT CCAAACCAAG GAGGAAAGGA TATCACGTAC GATGGTGAGC AAGGGCGAGG AGCTGTTCAC CGGGGTGGTG 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

CCCATCCTGG TCGAGCTGGA CGGCGACGTA AACGGCCACA AGTTCAGCGT GTCCGGCGAG GGCGAGGGCG ATGCCACCTA CGGCAAGCTG ACCCTGAAGT 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

TCATCTGCAC CACCGGCAAG CTGCCCGTGC CCTGGCCCAC CCTCGTGACC ACCTTGACCT ACGGCGTGCA GTGCTTCGCC CGCTACCCCG ACCACATGAA 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

GCAGCACGAC TTCTTCAAGT CCGCCATGCC CGAAGGCTAC GTCCAGGAGC GCACCATCTT CTTCAAGGAC GACGGCAACT ACAAGACCCG CGCCGAGGTG 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

AAGTTCGAGG GCGACACCCT GGTGAACCGC ATCGAGCTGA AGGGCATCGA CTTCAAGGAG GACGGCAACA TCCTGGGGCA CAAGCTGGAG TACAACTACA 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

ACAGCCACAA GGTCTATATC ACCGCCGACA AGCAGAAGAA CGGCATCAAG GTGAACTTCA AGACCCGCCA CAACATCGAG GACGGCAGCG TGCAGCTCGC 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

CGACCACTAC CAGCAGAACA CCCCCATCGG CGACGGCCCC GTGCTGCTGC CCGACAACCA CTACCTGAGC ACCCAGTCCG CCCTGAGCAA AGACCCCAAC 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

GAGAAGCGCG ATCACATGGT CCTGCTGGAG TTCGTGACCG CCGCCGGGAT CACTCTCGGC ATGGACGAGC TGTACAAGTA GCGGCCCAAT CTAGAGTCGG 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

GGCGGCCGGC CGCTTCGAGC AGACATGATA AGATACATTG ATGAGTTTGG ACAAACCACA ACTAGAATGC AGTGAAAAAA ATGCTTTATT TGTGAAATTT 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

GTGATGCTAT TGCTTTATTT GTAACCATTA TAAGCTGCAA TAAACAAGTT AACAACAACA ATTGCATTCA TTTTATGTTT CAGGTTCAGG GGGAGGTGTG 
^2601      ^2611      ^2621      ^2631      ^2641      ^2651      ^2661      ^2671      ^2681      ^2691      

GGAGGTTTTT TAAAGCAAGT AAAACCTCTA CAAATGTGGT AAAATCGATA AGGATCCGTC GACCGATGCC CTTGAGAGCC TTCAACCCAG TCAGCTCCTT 
^2701      ^2711      ^2721      ^2731      ^2741      ^2751      ^2761      ^2771      ^2781      ^2791      

CCGGTGGGCG CGGGGCATGA CTATCGTCGC CGCACTTATG ACTGTCTTCT TTATCATGCA ACTCGTAGGA CAGGTGCCGG CAGCGCTCTT CCGCTTCCTC 
^2801      ^2811      ^2821      ^2831      ^2841      ^2851      ^2861      ^2871      ^2881      ^2891      

GCTCACTGAC TCGCTGCGCT CGGTCGTTCG GCTGCGGCGA GCGGTATCAG CTCACTCAAA GGCGGTAATA CGGTTATCCA CAGAATCAGG GGATAACGCA 
^2901      ^2911      ^2921      ^2931      ^2941      ^2951      ^2961      ^2971      ^2981      ^2991      

GGAAAGAACA TGTGAGCAAA AGGCCAGCAA AAGGCCAGGA ACCGTAAAAA GGCCGCGTTG CTGGCGTTTT TCCATAGGCT CCGCCCCCCT GACGAGCATC 
^3001      ^3011      ^3021      ^3031      ^3041      ^3051      ^3061      ^3071      ^3081      ^3091      

ACAAAAATCG ACGCTCAAGT CAGAGGTGGC GAAACCCGAC AGGACTATAA AGATACCAGG CGTTTCCCCC TGGAAGCTCC CTCGTGCGCT CTCCTGTTCC 
^3101      ^3111      ^3121      ^3131      ^3141      ^3151      ^3161      ^3171      ^3181      ^3191      

GACCCTGCCG CTTACCGGAT ACCTGTCCGC CTTTCTCCCT TCGGGAAGCG TGGCGCTTTC TCATAGCTCA CGCTGTAGGT ATCTCAGTTC GGTGTAGGTC 
^3201      ^3211      ^3221      ^3231      ^3241      ^3251      ^3261      ^3271      ^3281      ^3291      

GTTCGCTCCA AGCTGGGCTG TGTGCACGAA CCCCCCGTTC AGCCCGACCG CTGCGCCTTA TCCGGTAACT ATCGTCTTGA GTCCAACCCG GTAAGACACG 
^3301      ^3311      ^3321      ^3331      ^3341      ^3351      ^3361      ^3371      ^3381      ^3391      

ACTTATCGCC ACTGGCAGCA GCCACTGGTA ACAGGATTAG CAGAGCGAGG TATGTAGGCG GTGCTACAGA GTTCTTGAAG TGGTGGCCTA ACTACGGCTA 
^3401      ^3411      ^3421      ^3431      ^3441      ^3451      ^3461      ^3471      ^3481      ^3491      

CACTAGAAGA ACAGTATTTG GTATCTGCGC TCTGCTGAAG CCAGTTACCT TCGGAAAAAG AGTTGGTAGC TCTTGATCCG GCAAACAAAC CACCGCTGGT 
^3501      ^3511      ^3521      ^3531      ^3541      ^3551      ^3561      ^3571      ^3581      ^3591      

AGCGGTGGTT TTTTTGTTTG CAAGCAGCAG ATTACGCGCA GAAAAAAAGG ATCTCAAGAA GATCCTTTGA TCTTTTCTAC GGGGTCTGAC GCTCAGTGGA 
^3601      ^3611      ^3621      ^3631      ^3641      ^3651      ^3661      ^3671      ^3681      ^3691      

ACGAAAACTC ACGTTAAGGG ATTTTGGTCA TGAGATTATC AAAAAGGATC TTCACCTAGA TCCTTTTAAA TTAAAAATGA AGTTTTAAAT CAATCTAAAG 
^3701      ^3711      ^3721      ^3731      ^3741      ^3751      ^3761      ^3771      ^3781      ^3791      

TATATATGAG TAAACTTGGT CTGACAGCGG CCGCAAATGC TAAACCACTG CAGTGGTTAC CAGTGCTTGA TCAGTGAGGC ACCGATCTCA GCGATCTGCC 
^3801      ^3811      ^3821      ^3831      ^3841      ^3851      ^3861      ^3871      ^3881      ^3891      

TATTTCGTTC GTCCATAGTG GCCTGACTCC CCGTCGTGTA GATCACTACG ATTCGTGAGG GCTTACCATC AGGCCCCAGC GCAGCAATGA TGCCGCGAGA 
^3901      ^3911      ^3921      ^3931      ^3941      ^3951      ^3961      ^3971      ^3981      ^3991      

GCCGCGTTCA CCGGCCCCCG ATTTGTCAGC AATGAACCAG CCAGCAGGGA GGGCCGAGCG AAGAAGTGGT CCTGCTACTT TGTCCGCCTC CATCCAGTCT 
^4001      ^4011      ^4021      ^4031      ^4041      ^4051      ^4061      ^4071      ^4081      ^4091      

ATGAGCTGCT GTCGTGATGC TAGAGTAAGA AGTTCGCCAG TGAGTAGTTT CCGAAGAGTT GTGGCCATTG CTACTGGCAT CGTGGTATCA CGCTCGTCGT 
^4101      ^4111      ^4121      ^4131      ^4141      ^4151      ^4161      ^4171      ^4181      ^4191      

TCGGTATGGC TTCGTTCAAC TCTGGTTCCC AGCGGTCAAG CCGGGTCACA TGATCACCCA TATTATGAAG AAATGCAGTC AGCTCCTTAG GGCCTCCGAT 
^4201      ^4211      ^4221      ^4231      ^4241      ^4251      ^4261      ^4271      ^4281      ^4291      

CGTTGTCAGA AGTAAGTTGG CCGCGGTGTT GTCGCTCATG GTAATGGCAG CACTACACAA TTCTCTTACC GTCATGCCAT CCGTAAGATG CTTTTCCGTG 
^4301      ^4311      ^4321      ^4331      ^4341      ^4351      ^4361      ^4371      ^4381      ^4391      

ACCGGCGAGT ACTCAACCAA GTCGTTTTGT GAGTAGTGTA TACGGCGACC AAGCTGCTCT TGCCCGGCGT CTATACGGGA CAACACCGCG CCACATAGCA 
^4401      ^4411      ^4421      ^4431      ^4441      ^4451      ^4461      ^4471      ^4481      ^4491      

GTACTTTGAA AGTGCTCATC ATCGGGAATC GTTCTTCGGG GCGGAAAGAC TCAAGGATCT TGCCGCTATT GAGATCCAGT TCGATATAGC CCACTCTTGC 
^4501      ^4511      ^4521      ^4531      ^4541      ^4551      ^4561      ^4571      ^4581      ^4591      

ACCCAGTTGA TCTTCAGCAT CTTTTACTTT CACCAGCGTT TCGGGGTGTG CAAAAACAGG CAAGCAAAAT GCCGCAAAGA AGGGAATGAG TGCGACACGA 
^4601      ^4611      ^4621      ^4631      ^4641      ^4651      ^4661      ^4671      ^4681      ^4691      

AAATGTTGGA TGCTCATACT CGTCCTTTTT CAATATTATT GAAGCATTTA TCAGGGTTAC TAGTACGTCT CTCAAGGATA AGTAAGTAAT ATTAAGGTAC 
^4701      ^4711      ^4721      ^4731      ^4741      ^4751      ^4761      ^4771      ^4781      ^4791      

GGGAGGTATT GGACAGGCCG CAATAAAATA TCTTTATTTT CATTACATCT GTGTGTTGGT TTTTTGTGTG AATCGATAGT ACTAACATAC GCTCTCCATC 
^4801      ^4811      ^4821      ^4831      ^4841      ^4851      ^4861      ^4871      ^4881      ^4891      

AAAACAAAAC GAAACAAAAC AAACTAGCAA AATAGGCTGT CCCCAGTGCA AGTGCAGGTG CCAGAACATT TCTCTGGCCT AACTGGCC
^4901      ^4911      ^4921      ^4931      ^4941      ^4951      ^4961      ^4971      ^4981

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:
GLuc

The description of the protein encoded in this ORF:
Gaussia 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):

91-648

ORF

ORF position:

2

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):

1028-1747

Remarks:

In addition to the unspliced message 4 alternatively spliced products were reported in HeLa cells transiently
transfected by this plasmid. Although the authors intended to use a plasmid which would be depleted of
5'-splice donor sites (Supplementary Figure 6) still several 5'-splice donor sites remained in GLuc coding
region and even in the eIF4G sequence itself. Please refer to the Supplementary Figure 7 in Baranick et al.
(2008).

The sequences around the 5'-splice donor were:

TGCTGGTGGGgtagggatga (in the eIF4G sequence itself)

GGGCCCCAGGgtgtgcagcc (in the eIF4G sequence itself, intron emphasized in lowercase letters of this IRESite
record)

CCAGGGCCAAgtcgataaga (in GLuc coding region, C-terminus)

ACCTTCGCAAgcaagatcca (in GLuc coding region, C-terminus, also functional in XIAP-based plasmid IRESite_Id:563)

The 3'-splice acceptor region within eIF4G was: ttatagctttctttccccagAT

Comments from IRESite curator:

The first splice variant has intercistronic region of size 122bp (instead of 379bp as the unspliced message)
while both reported ORFs intact.

The second splice variant has intercistronic region of size 88bp (instead of 379bp as the unspliced message)
while both reported ORFs intact.

The third splice variant has the first cistron merged in another frame to the second cistron, quite
surprisingly the fusion protein extends even behind the GFP protein ORF by 17 bp. Scanning ribosome would have
to bypass AUGs of 6-10 ORFs 538-720bp to synthesize GFP protein (of these are 2 uORFs).

The fourth splice variant has modified C-terminal region of the first cistron spanning the eIF4GI 5'-UTR
insert sequence and hits a STOP codon in the very beginning of GFP ORF. However, in another ORF there could be
produced GFP protein in its original reading frame with extra 26 aminoacid residues fused to it N-terminus.
And, obviously unaltered GFP protein could be produced as well. Scanning ribosome would have to bypass AUGs of
6-9 ORFs over a distance of 538-644bp to synthesize GFP protein (of these are 3 uORFs).

Citations:

Baranick B. T., Lemp N. A., Nagashima J., Hiraoka K., Kasahara N., Logg C. R. (2008) Splicing mediates the activity of four putative cellular internal ribosome entry sites. Proc. Natl. Acad. Sci. U. S. A. 105(12):4733-4738

IRESs:

IRES:

Version: `2`	Last change: `2009-09-02 22:13:19`
Originaly submitted by: Martin Mokrejš	Reviewed by: Martin Mokrejš
The IRES name: `eIF4G_-368_-12`	The functional status of IRES: `functional`

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

665-1021

How IRES boundaries were determined:
experimentally_determined

5'-end of IRES relative to last base of the STOP codon of the upstream ORF: `17`	3'-end of IRES relative to last base of the STOP codon of the upstream ORF: `373`
5'-end of IRES relative to first base of the START codon of the downstream ORF: `-363`	3'-end of IRES relative to first base of the START codon of the downstream ORF: `-7`

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

TCTAGATGGG GGTCCTGGGC CCCAGGgtgt gcagccactg acttggggac tgctggtggg gtagggatga gggagggagg ggcattgtga tgtacagggc 
^665       ^675       ^685       ^695       ^705       ^715       ^725       ^735       ^745       ^755       

tgctctgtga gatcaagggt ctcttaaggg tgggagctgg ggcagggact acgagagcag ccagatgggc tgaaagtgga actcaagggg tttctggcac 
^765       ^775       ^785       ^795       ^805       ^815       ^825       ^835       ^845       ^855       

ctacctacct gcttcccgct ggggggtggg gagttggccc agagtcttaa gattggggca gggtggagag gtgggctctt cctgcttccc actcatctta 
^865       ^875       ^885       ^895       ^905       ^915       ^925       ^935       ^945       ^955       

tagctttctt tccccagATC CGAATTCGAG ATCCAAACCA AGGAGGAAAG GATATCA
^965       ^975       ^985       ^995       ^1005      ^1015

Citations:

The translation experiments:

Translation results:

IRESite Translation Id: 601

Version: `1`	Last change: `2009-07-16 14:37:08`
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:

21.700

Name of IRES used as the positive control:

EMCV-R_-541_-7

Name of the plasmid used as the positive control. `pGLuc-EMCV-GFP`	Name of the plasmid used as the negative control. `pGLuc-GFP`
`IRESite Id` of the plasmid used as positive control. `557`	`IRESite Id` of the plasmid used as negative control. `556`
The relative translation efficiency in % of the positive control: `100.000`	The relative translation efficiency in % of the negative control: `10.800`
The size (length) of intercistronic region in the positive control: `565`	The size (length) of intercistronic region in the negative control: `17`

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:

Data from Figure 1B.

Citations:

Translation results:

IRESite Translation Id: 602

Version: `1`	Last change: `2009-07-16 14:37:08`
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 NIH/3T3 (ATCC CRL-1658)

The temperature (in degrees of Celsia):
37

The relative translation efficiency in % of this IRES:

16.600

Name of IRES used as the positive control:

EMCV-R_-541_-7

Name of the plasmid used as the positive control. `pGLuc-EMCV-GFP`	Name of the plasmid used as the negative control. `pGLuc-GFP`
`IRESite Id` of the plasmid used as positive control. `557`	`IRESite Id` of the plasmid used as negative control. `556`
The relative translation efficiency in % of the positive control: `100.000`	The relative translation efficiency in % of the negative control: `10.900`
The size (length) of intercistronic region in the positive control: `565`	The size (length) of intercistronic region in the negative control: `17`

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:

Data from Figure 1B.

Citations:

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