IRESite_Id:90 characteristics of HCV_type_1a IRES studied in pFGAL4-HCV1 plasmid

IRESite: The database of experimentally verified IRES structures

The nucleic acid data:

IRESite Id: `90`	Version: `8`
Originaly submitted by: Tomáš Mašek	Submission date: `2005-12-15 00:00:00`
Reviewed by: Tomáš Mašek	Last change: `2008-07-04 12:34:26`

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:

mRNA produced by bicistronic plasmid pFGAL4-HCV1 which comprises luciferase and Gal4 genes as the first and
the second cistron, respectively.The full-length 5'-UTR of HCV genomic RNA together with the first 45 nt of a
viral polyprotein gene was inserted in frame into the intercistronic region of the reporter pFGAL4h vector,
thus making the N-terminal fusion of the first 15 amino acids residues from the HCV polyprotein with the yeast
transcriptional activator Gal4p.

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

ATGGAAGACG CCAAAAACAT AAAGAAAGGC CCGGCGCCAT TCTATCCGCT GGAAGATGGA ACCGCTGGAG AGCAACTGCA TAAGGCTATG AAGAGATACG 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

CCCTGGTTCC TGGAACAATT GCTTTTACAG ATGCACATAT CGAGGTGGAC ATCACTTACG CTGAGTACTT CGAAATGTCC GTTCGGTTGG CAGAAGCTAT 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

GAAACGATAT GGGCTGAATA CAAATCACAG AATCGTCGTA TGCAGTGAAA ACTCTCTTCA ATTCTTTATG CCGGTGTTGG GCGCGTTATT TATCGGAGTT 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

GCAGTTGCGC CCGCGAACGA CATTTATAAT GAACGTGAAT TGCTCAACAG TATGGGCATT TCGCAGCCTA CCGTGGTGTT CGTTTCCAAA AAGGGGTTGC 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

AAAAAATTTT GAACGTGCAA AAAAAGCTCC CAATCATCCA AAAAATTATT ATCATGGATT CTAAAACGGA TTACCAGGGA TTTCAGTCGA TGTACACGTT 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

CGTCACATCT CATCTACCTC CCGGTTTTAA TGAATACGAT TTTGTGCCAG AGTCCTTCGA TAGGGACAAG ACAATTGCAC TGATCATGAA CTCCTCTGGA 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

TCTACTGGTC TGCCTAAAGG TGTCGCTCTG CCTCATAGAA CTGCCTGCGT GAGATTCTCG CATGCCAGAG ATCCTATTTT TGGCAATCAA ATCATTCCGG 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

ATACTGCGAT TTTAAGTGTT GTTCCATTCC ATCACGGTTT TGGAATGTTT ACTACACTCG GATATTTGAT ATGTGGATTT CGAGTCGTCT TAATGTATAG 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

ATTTGAAGAA GAGCTGTTTC TGAGGAGCCT TCAGGATTAC AAGATTCAAA GTGCGCTGCT GGTGCCAACC CTATTCTCCT TCTTCGCCAA AAGCACTCTG 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

ATTGACAAAT ACGATTTATC TAATTTACAC GAAATTGCTT CTGGTGGCGC TCCCCTCTCT AAGGAAGTCG GGGAAGCGGT TGCCAAGAGG TTCCATCTGC 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

CAGGTATCAG GCAAGGATAT GGGCTCACTG AGACTACATC AGCTATTCTG ATTACACCCG AGGGGGATGA TAAACCGGGC GCGGTCGGTA AAGTTGTTCC 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

ATTTTTTGAA GCGAAGGTTG TGGATCTGGA TACCGGGAAA ACGCTGGGCG TTAATCAAAG AGGCGAACTG TGTGTGAGAG GTCCTATGAT TATGTCCGGT 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

TATGTAAACA ATCCGGAAGC GACCAACGCC TTGATTGACA AGGATGGATG GCTACATTCT GGAGACATAG CTTACTGGGA CGAAGACGAA CACTTCTTCA 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

TCGTTGACCG CCTGAAGTCT CTGATTAAGT ACAAAGGCTA TCAGGTGGCT CCCGCTGAAT TGGAATCCAT CTTGCTCCAA CACCCCAACA TCTTCGACGC 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

AGGTGTCGCA GGTCTTCCCG ACGATGACGC CGGTGAACTT CCCGCCGCCG TTGTTGTTTT GGAGCACGGA AAGACGATGA CGGAAAAAGA GATCGTGGAT 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

TACGTCGCCA GTCAAGTAAC AACCGCGAAA AAGTTGCGCG GAGGAGTTGT GTTTGTGGAC GAAGTACCGA AAGGTCTTAC CGGAAAACTC GACGCAAGAA 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

AAATCAGAGA GATCCTCATA AAGGCCAAGA AGGGCGGAAA GATCGCCGTG TAATTCTAGC CTAGGATCCG CCAGCCCCCT GATGGGGGCG ACACTCCACC 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

ATGAATCACT CCCCTGTGAG GAACTACTGT CTTCACGCAG AAAGCGTCTA GCCATGGCGT TAGTATGAGT GTCGTGCAGC CTCCAGGACC CCCCCTCCCG 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

GGAGAGCCAT AGTGGTCTGC GGAACCGGTG AGTACACCGG AATTGCCAGG ACGACCGGGT CCTTTCTTGG ATAAACCCGC TCAATGCCTG GAGATTTGGG 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

CGTGCCCCCG CAAGACTGCT AGCCGAGTAG TGTTGGGTCG CGAAAGGCCT TGTGGTACTG CCTGATAGGG TGCTTGCGAG TGCCCCGGGA GGTCTCGTAG 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

ACCGTGCACC ATGAGCACGA ATCCTAAACC TCAAAGAAAA ACCAAACGTA ACACGAATTC AAAAATGAAG CTACTGTCTT CTATCGAACA AGCATGCGAT 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

ATTTGCCGAC TTAAAAAGCT CAAGTGCTCC AAAGAAAAAC CGAAGTGCGC CAAGTGTCTG AAGAACAACT GGGAGTGTCG CTACTCTCCC AAAACCAAAA 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

GGTCTCCGCT GACTAGGGCA CATCTGACAG AAGTGGAATC AAGGCTAGAA AGACTGGAAC AGCTATTTCT ACTGATTTTT CCTCGAGAAG ACCTTGACAT 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

GATTTTGAAA ATGGATTCTT TACAGGATAT AAAAGCATTG TTAACAGGAT TATTTGTACA AGATAATGTG AATAAAGATG CCGTCACAGA TAGATTGGCT 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

TCAGTGGAGA CTGATATGCC TCTAACATTG AGACAGCATA GAATAAGTGC GACATCATCA TCGGAAGAGA GTAGTAACAA AGGTCAAAGA CAGTTGACTG 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

TATCGATTGA CTCGGCAGCT CATCATGATA ACTCCACAAT TCCGTTGGAT TTTATGCCCA GGGATGCTCT TCATGGATTT GATTGGTCTG AAGAGGATGA 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

CATGTCGGAT GGCTTGCCCT TCCTGAAAAC GGACCCCAAC AATAATGGGT TCTTTGGCGA CGGTTCTCTC TTATGTATTC TTCGATCTAT TGGCTTTAAA 
^2601      ^2611      ^2621      ^2631      ^2641      ^2651      ^2661      ^2671      ^2681      ^2691      

CCGGAAAATT ACACGAACTC TAACGTTAAC AGGCTCCCGA CCATGATTAC GGATAGATAC ACGTTGGCTT CTAGATCCAC AACATCCCGT TTACTTCAAA 
^2701      ^2711      ^2721      ^2731      ^2741      ^2751      ^2761      ^2771      ^2781      ^2791      

GTTATCTCAA TAATTTTCAC CCCTACTGCC CTATCGTGCA CTCACCGACG CTAATGATGT TGTATAATAA CCAGATTGAA ATCGCGTCGA AGGATCAATG 
^2801      ^2811      ^2821      ^2831      ^2841      ^2851      ^2861      ^2871      ^2881      ^2891      

GCAAATCCTT TTTAACTGCA TATTAGCCAT TGGAGCCTGG TGTATAGAGG GGGAATCTAC TGATATAGAT GTTTTTTACT ATCAAAATGC TAAATCTCAT 
^2901      ^2911      ^2921      ^2931      ^2941      ^2951      ^2961      ^2971      ^2981      ^2991      

TTGACGAGCA AGGTCTTCGA GTCAGGTTCC ATAATTTTGG TGACAGCCCT ACATCTTCTG TCGCGATATA CACAGTGGAG GCAGAAAACA AATACTAGCT 
^3001      ^3011      ^3021      ^3031      ^3041      ^3051      ^3061      ^3071      ^3081      ^3091      

ATAATTTTCA CAGCTTTTCC ATAAGAATGG CCATATCATT GGGCTTGAAT AGGGACCTCC CCTCGTCCTT CAGTGATAGC AGCATTCTGG AACAAAGACG 
^3101      ^3111      ^3121      ^3131      ^3141      ^3151      ^3161      ^3171      ^3181      ^3191      

CCGAATTTGG TGGTCTGTCT ACTCTTGGGA GATCCAATTG TCCCTGCTTT ATGGTCGATC CATCCAGCTT TCTCAGAATA CAATCTCCTT CCCTTCTTCT 
^3201      ^3211      ^3221      ^3231      ^3241      ^3251      ^3261      ^3271      ^3281      ^3291      

GTCGACGATG TGCAGCGTAC CACAACAGGT CCCACCATAT ATCATGGCAT CATTGAAACA GCAAGGCTCT TACAAGTTTT CACAAAAATC TATGAACTAG 
^3301      ^3311      ^3321      ^3331      ^3341      ^3351      ^3361      ^3371      ^3381      ^3391      

ACAAAACAGT AACTGCAGAA AAAAGTCCTA TATGTGCAAA AAAATGCTTG ATGATTTGTA ATGAGATTGA GGAGGTTTCG AGACAGGCAC CAAAGTTTTT 
^3401      ^3411      ^3421      ^3431      ^3441      ^3451      ^3461      ^3471      ^3481      ^3491      

ACAAATGGAT ATTTCCACCA CCGCTCTAAC CAATTTGTTG AAGGAACACC CTTGGCTATC CTTTACAAGA TTCGAACTGA AGTGGAAACA GTTGTCTCTT 
^3501      ^3511      ^3521      ^3531      ^3541      ^3551      ^3561      ^3571      ^3581      ^3591      

ATCATTTATG TATTAAGAGA TTTTTTCACT AATTTTACCC AGAAAAAGTC ACAACTAGAA CAGGATCAAA ATGATCATCA AAGTTATGAA GTTAAACGAT 
^3601      ^3611      ^3621      ^3631      ^3641      ^3651      ^3661      ^3671      ^3681      ^3691      

GCTCCATCAT GTTAAGCGAT GCAGCACAAA GAACTGTTAT GTCTGTAAGT AGCTATATGG ACAATCATAA TGTCACCCCA TATTTTGCCT GGAATTGTTC 
^3701      ^3711      ^3721      ^3731      ^3741      ^3751      ^3761      ^3771      ^3781      ^3791      

TTATTACTTG TTCAATGCAG TCCTAGTACC CATAAAGACT CTACTCTCAA ACTCAAAATC GAATGCTGAG AATAACGAGA CCGCACAATT ATTACAACAA 
^3801      ^3811      ^3821      ^3831      ^3841      ^3851      ^3861      ^3871      ^3881      ^3891      

ATTAACACTG TTCTGATGCT ATTAAAAAAA CTGGCCACTT TTAAAATCCA GACTTGTGAA AAATACATTC AAGTACTGGA AGAGGTATGT GCGCCGTTTC 
^3901      ^3911      ^3921      ^3931      ^3941      ^3951      ^3961      ^3971      ^3981      ^3991      

TGTTATCACA GTGTGCAATC CCATTACCGC ATATCAGTTA TAACAATAGT AATGGTAGCG CCATTAAAAA TATTGTCGGT TCTGCAACTA TCGCCCAATA 
^4001      ^4011      ^4021      ^4031      ^4041      ^4051      ^4061      ^4071      ^4081      ^4091      

CCCTACTCTT CCGGAGGAAA ATGTCAACAA TATCAGTGTT AAATATGTTT CTCCTGGCTC AGTAGGGCCT TCACCTGTGC CATTGAAATC AGGAGCAAGT 
^4101      ^4111      ^4121      ^4131      ^4141      ^4151      ^4161      ^4171      ^4181      ^4191      

TTCAGTGATC TAGTCAAGCT GTTATCTAAC CGTCCACCCT CTCGTAACTC TCCAGTGACA ATACCAAGAA GCACACCTTC GCATCGCTCA GTCACGCCTT 
^4201      ^4211      ^4221      ^4231      ^4241      ^4251      ^4261      ^4271      ^4281      ^4291      

TTCTAGGGCA ACAGCAACAG CTGCAATCAT TAGTGCCACT GACCCCGTCT GCTTTGTTTG GTGGCGCCAA TTTTAATCAA AGTGGGAATA TTGCTGATAG 
^4301      ^4311      ^4321      ^4331      ^4341      ^4351      ^4361      ^4371      ^4381      ^4391      

CTCATTGTCC TTCACTTTCA CTAACAGTAG CAACGGTCCG AACCTCATAA CAACTCAAAC AAATTCTCAA GCGCTTTCAC AACCAATTGC CTCCTCTAAC 
^4401      ^4411      ^4421      ^4431      ^4441      ^4451      ^4461      ^4471      ^4481      ^4491      

GTTCATGATA ACTTCATGAA TAATGAAATC ACGGCTAGTA AAATTGATGA TGGTAATAAT TCAAAACCAC TGTCACCTGG TTGGACGGAC CAAACTGCGT 
^4501      ^4511      ^4521      ^4531      ^4541      ^4551      ^4561      ^4571      ^4581      ^4591      

ATAACGCGTT TGGAATCACT ACAGGGATGT TTAATACCAC TACAATGGAT GATGTATATA ACTATCTATT CGATGATGAA GATACCCCAC CAAACCCAAA 
^4601      ^4611      ^4621      ^4631      ^4641      ^4651      ^4661      ^4671      ^4681      ^4691      

AAAAGAGTAA
^4701

Credibility of mRNA sequence:

end-to-end_sequence_completely_same_as_in_the_experiment

The name of the plasmid:
pFGAL4-HCV1 The name of the promoter used to express this mRNA:
TPI

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

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:
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:

Saccharomyces cerevisiae pJ69a

The abbreviated name of the donor gene or virus from which this IRES was excised and inserted into the plasmid:
HCV1a

The origin of IRES in the plasmid:

viral

The donor organism of the IRES segment:

Hepatitis C virus type 1a

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

GAATTAATTC ACCATGGAAG ACGCCAAAAA CATAAAGAAA GGCCCGGCGC CATTCTATCC GCTGGAAGAT GGAACCGCTG GAGAGCAACT GCATAAGGCT 
^1         ^11        ^21        ^31        ^41        ^51        ^61        ^71        ^81        ^91        

ATGAAGAGAT ACGCCCTGGT TCCTGGAACA ATTGCTTTTA CAGATGCACA TATCGAGGTG GACATCACTT ACGCTGAGTA CTTCGAAATG TCCGTTCGGT 
^101       ^111       ^121       ^131       ^141       ^151       ^161       ^171       ^181       ^191       

TGGCAGAAGC TATGAAACGA TATGGGCTGA ATACAAATCA CAGAATCGTC GTATGCAGTG AAAACTCTCT TCAATTCTTT ATGCCGGTGT TGGGCGCGTT 
^201       ^211       ^221       ^231       ^241       ^251       ^261       ^271       ^281       ^291       

ATTTATCGGA GTTGCAGTTG CGCCCGCGAA CGACATTTAT AATGAACGTG AATTGCTCAA CAGTATGGGC ATTTCGCAGC CTACCGTGGT GTTCGTTTCC 
^301       ^311       ^321       ^331       ^341       ^351       ^361       ^371       ^381       ^391       

AAAAAGGGGT TGCAAAAAAT TTTGAACGTG CAAAAAAAGC TCCCAATCAT CCAAAAAATT ATTATCATGG ATTCTAAAAC GGATTACCAG GGATTTCAGT 
^401       ^411       ^421       ^431       ^441       ^451       ^461       ^471       ^481       ^491       

CGATGTACAC GTTCGTCACA TCTCATCTAC CTCCCGGTTT TAATGAATAC GATTTTGTGC CAGAGTCCTT CGATAGGGAC AAGACAATTG CACTGATCAT 
^501       ^511       ^521       ^531       ^541       ^551       ^561       ^571       ^581       ^591       

GAACTCCTCT GGATCTACTG GTCTGCCTAA AGGTGTCGCT CTGCCTCATA GAACTGCCTG CGTGAGATTC TCGCATGCCA GAGATCCTAT TTTTGGCAAT 
^601       ^611       ^621       ^631       ^641       ^651       ^661       ^671       ^681       ^691       

CAAATCATTC CGGATACTGC GATTTTAAGT GTTGTTCCAT TCCATCACGG TTTTGGAATG TTTACTACAC TCGGATATTT GATATGTGGA TTTCGAGTCG 
^701       ^711       ^721       ^731       ^741       ^751       ^761       ^771       ^781       ^791       

TCTTAATGTA TAGATTTGAA GAAGAGCTGT TTCTGAGGAG CCTTCAGGAT TACAAGATTC AAAGTGCGCT GCTGGTGCCA ACCCTATTCT CCTTCTTCGC 
^801       ^811       ^821       ^831       ^841       ^851       ^861       ^871       ^881       ^891       

CAAAAGCACT CTGATTGACA AATACGATTT ATCTAATTTA CACGAAATTG CTTCTGGTGG CGCTCCCCTC TCTAAGGAAG TCGGGGAAGC GGTTGCCAAG 
^901       ^911       ^921       ^931       ^941       ^951       ^961       ^971       ^981       ^991       

AGGTTCCATC TGCCAGGTAT CAGGCAAGGA TATGGGCTCA CTGAGACTAC ATCAGCTATT CTGATTACAC CCGAGGGGGA TGATAAACCG GGCGCGGTCG 
^1001      ^1011      ^1021      ^1031      ^1041      ^1051      ^1061      ^1071      ^1081      ^1091      

GTAAAGTTGT TCCATTTTTT GAAGCGAAGG TTGTGGATCT GGATACCGGG AAAACGCTGG GCGTTAATCA AAGAGGCGAA CTGTGTGTGA GAGGTCCTAT 
^1101      ^1111      ^1121      ^1131      ^1141      ^1151      ^1161      ^1171      ^1181      ^1191      

GATTATGTCC GGTTATGTAA ACAATCCGGA AGCGACCAAC GCCTTGATTG ACAAGGATGG ATGGCTACAT TCTGGAGACA TAGCTTACTG GGACGAAGAC 
^1201      ^1211      ^1221      ^1231      ^1241      ^1251      ^1261      ^1271      ^1281      ^1291      

GAACACTTCT TCATCGTTGA CCGCCTGAAG TCTCTGATTA AGTACAAAGG CTATCAGGTG GCTCCCGCTG AATTGGAATC CATCTTGCTC CAACACCCCA 
^1301      ^1311      ^1321      ^1331      ^1341      ^1351      ^1361      ^1371      ^1381      ^1391      

ACATCTTCGA CGCAGGTGTC GCAGGTCTTC CCGACGATGA CGCCGGTGAA CTTCCCGCCG CCGTTGTTGT TTTGGAGCAC GGAAAGACGA TGACGGAAAA 
^1401      ^1411      ^1421      ^1431      ^1441      ^1451      ^1461      ^1471      ^1481      ^1491      

AGAGATCGTG GATTACGTCG CCAGTCAAGT AACAACCGCG AAAAAGTTGC GCGGAGGAGT TGTGTTTGTG GACGAAGTAC CGAAAGGTCT TACCGGAAAA 
^1501      ^1511      ^1521      ^1531      ^1541      ^1551      ^1561      ^1571      ^1581      ^1591      

CTCGACGCAA GAAAAATCAG AGAGATCCTC ATAAAGGCCA AGAAGGGCGG AAAGATCGCC GTGTAATTCT AGCCTAGGAT CCGCCAGCCC CCTGATGGGG 
^1601      ^1611      ^1621      ^1631      ^1641      ^1651      ^1661      ^1671      ^1681      ^1691      

GCGACACTCC ACCATGAATC ACTCCCCTGT GAGGAACTAC TGTCTTCACG CAGAAAGCGT CTAGCCATGG CGTTAGTATG AGTGTCGTGC AGCCTCCAGG 
^1701      ^1711      ^1721      ^1731      ^1741      ^1751      ^1761      ^1771      ^1781      ^1791      

ACCCCCCCTC CCGGGAGAGC CATAGTGGTC TGCGGAACCG GTGAGTACAC CGGAATTGCC AGGACGACCG GGTCCTTTCT TGGATAAACC CGCTCAATGC 
^1801      ^1811      ^1821      ^1831      ^1841      ^1851      ^1861      ^1871      ^1881      ^1891      

CTGGAGATTT GGGCGTGCCC CCGCAAGACT GCTAGCCGAG TAGTGTTGGG TCGCGAAAGG CCTTGTGGTA CTGCCTGATA GGGTGCTTGC GAGTGCCCCG 
^1901      ^1911      ^1921      ^1931      ^1941      ^1951      ^1961      ^1971      ^1981      ^1991      

GGAGGTCTCG TAGACCGTGC ACCATGAGCA CGAATCCTAA ACCTCAAAGA AAAACCAAAC GTAACACGAA TTCAAAAATG AAGCTACTGT CTTCTATCGA 
^2001      ^2011      ^2021      ^2031      ^2041      ^2051      ^2061      ^2071      ^2081      ^2091      

ACAAGCATGC GATATTTGCC GACTTAAAAA GCTCAAGTGC TCCAAAGAAA AACCGAAGTG CGCCAAGTGT CTGAAGAACA ACTGGGAGTG TCGCTACTCT 
^2101      ^2111      ^2121      ^2131      ^2141      ^2151      ^2161      ^2171      ^2181      ^2191      

CCCAAAACCA AAAGGTCTCC GCTGACTAGG GCACATCTGA CAGAAGTGGA ATCAAGGCTA GAAAGACTGG AACAGCTATT TCTACTGATT TTTCCTCGAG 
^2201      ^2211      ^2221      ^2231      ^2241      ^2251      ^2261      ^2271      ^2281      ^2291      

AAGACCTTGA CATGATTTTG AAAATGGATT CTTTACAGGA TATAAAAGCA TTGTTAACAG GATTATTTGT ACAAGATAAT GTGAATAAAG ATGCCGTCAC 
^2301      ^2311      ^2321      ^2331      ^2341      ^2351      ^2361      ^2371      ^2381      ^2391      

AGATAGATTG GCTTCAGTGG AGACTGATAT GCCTCTAACA TTGAGACAGC ATAGAATAAG TGCGACATCA TCATCGGAAG AGAGTAGTAA CAAAGGTCAA 
^2401      ^2411      ^2421      ^2431      ^2441      ^2451      ^2461      ^2471      ^2481      ^2491      

AGACAGTTGA CTGTATCGAT TGACTCGGCA GCTCATCATG ATAACTCCAC AATTCCGTTG GATTTTATGC CCAGGGATGC TCTTCATGGA TTTGATTGGT 
^2501      ^2511      ^2521      ^2531      ^2541      ^2551      ^2561      ^2571      ^2581      ^2591      

CTGAAGAGGA TGACATGTCG GATGGCTTGC CCTTCCTGAA AACGGACCCC AACAATAATG GGTTCTTTGG CGACGGTTCT CTCTTATGTA TTCTTCGATC 
^2601      ^2611      ^2621      ^2631      ^2641      ^2651      ^2661      ^2671      ^2681      ^2691      

TATTGGCTTT AAACCGGAAA ATTACACGAA CTCTAACGTT AACAGGCTCC CGACCATGAT TACGGATAGA TACACGTTGG CTTCTAGATC CACAACATCC 
^2701      ^2711      ^2721      ^2731      ^2741      ^2751      ^2761      ^2771      ^2781      ^2791      

CGTTTACTTC AAAGTTATCT CAATAATTTT CACCCCTACT GCCCTATCGT GCACTCACCG ACGCTAATGA TGTTGTATAA TAACCAGATT GAAATCGCGT 
^2801      ^2811      ^2821      ^2831      ^2841      ^2851      ^2861      ^2871      ^2881      ^2891      

CGAAGGATCA ATGGCAAATC CTTTTTAACT GCATATTAGC CATTGGAGCC TGGTGTATAG AGGGGGAATC TACTGATATA GATGTTTTTT ACTATCAAAA 
^2901      ^2911      ^2921      ^2931      ^2941      ^2951      ^2961      ^2971      ^2981      ^2991      

TGCTAAATCT CATTTGACGA GCAAGGTCTT CGAGTCAGGT TCCATAATTT TGGTGACAGC CCTACATCTT CTGTCGCGAT ATACACAGTG GAGGCAGAAA 
^3001      ^3011      ^3021      ^3031      ^3041      ^3051      ^3061      ^3071      ^3081      ^3091      

ACAAATACTA GCTATAATTT TCACAGCTTT TCCATAAGAA TGGCCATATC ATTGGGCTTG AATAGGGACC TCCCCTCGTC CTTCAGTGAT AGCAGCATTC 
^3101      ^3111      ^3121      ^3131      ^3141      ^3151      ^3161      ^3171      ^3181      ^3191      

TGGAACAAAG ACGCCGAATT TGGTGGTCTG TCTACTCTTG GGAGATCCAA TTGTCCCTGC TTTATGGTCG ATCCATCCAG CTTTCTCAGA ATACAATCTC 
^3201      ^3211      ^3221      ^3231      ^3241      ^3251      ^3261      ^3271      ^3281      ^3291      

CTTCCCTTCT TCTGTCGACG ATGTGCAGCG TACCACAACA GGTCCCACCA TATATCATGG CATCATTGAA ACAGCAAGGC TCTTACAAGT TTTCACAAAA 
^3301      ^3311      ^3321      ^3331      ^3341      ^3351      ^3361      ^3371      ^3381      ^3391      

ATCTATGAAC TAGACAAAAC AGTAACTGCA GAAAAAAGTC CTATATGTGC AAAAAAATGC TTGATGATTT GTAATGAGAT TGAGGAGGTT TCGAGACAGG 
^3401      ^3411      ^3421      ^3431      ^3441      ^3451      ^3461      ^3471      ^3481      ^3491      

CACCAAAGTT TTTACAAATG GATATTTCCA CCACCGCTCT AACCAATTTG TTGAAGGAAC ACCCTTGGCT ATCCTTTACA AGATTCGAAC TGAAGTGGAA 
^3501      ^3511      ^3521      ^3531      ^3541      ^3551      ^3561      ^3571      ^3581      ^3591      

ACAGTTGTCT CTTATCATTT ATGTATTAAG AGATTTTTTC ACTAATTTTA CCCAGAAAAA GTCACAACTA GAACAGGATC AAAATGATCA TCAAAGTTAT 
^3601      ^3611      ^3621      ^3631      ^3641      ^3651      ^3661      ^3671      ^3681      ^3691      

GAAGTTAAAC GATGCTCCAT CATGTTAAGC GATGCAGCAC AAAGAACTGT TATGTCTGTA AGTAGCTATA TGGACAATCA TAATGTCACC CCATATTTTG 
^3701      ^3711      ^3721      ^3731      ^3741      ^3751      ^3761      ^3771      ^3781      ^3791      

CCTGGAATTG TTCTTATTAC TTGTTCAATG CAGTCCTAGT ACCCATAAAG ACTCTACTCT CAAACTCAAA ATCGAATGCT GAGAATAACG AGACCGCACA 
^3801      ^3811      ^3821      ^3831      ^3841      ^3851      ^3861      ^3871      ^3881      ^3891      

ATTATTACAA CAAATTAACA CTGTTCTGAT GCTATTAAAA AAACTGGCCA CTTTTAAAAT CCAGACTTGT GAAAAATACA TTCAAGTACT GGAAGAGGTA 
^3901      ^3911      ^3921      ^3931      ^3941      ^3951      ^3961      ^3971      ^3981      ^3991      

TGTGCGCCGT TTCTGTTATC ACAGTGTGCA ATCCCATTAC CGCATATCAG TTATAACAAT AGTAATGGTA GCGCCATTAA AAATATTGTC GGTTCTGCAA 
^4001      ^4011      ^4021      ^4031      ^4041      ^4051      ^4061      ^4071      ^4081      ^4091      

CTATCGCCCA ATACCCTACT CTTCCGGAGG AAAATGTCAA CAATATCAGT GTTAAATATG TTTCTCCTGG CTCAGTAGGG CCTTCACCTG TGCCATTGAA 
^4101      ^4111      ^4121      ^4131      ^4141      ^4151      ^4161      ^4171      ^4181      ^4191      

ATCAGGAGCA AGTTTCAGTG ATCTAGTCAA GCTGTTATCT AACCGTCCAC CCTCTCGTAA CTCTCCAGTG ACAATACCAA GAAGCACACC TTCGCATCGC 
^4201      ^4211      ^4221      ^4231      ^4241      ^4251      ^4261      ^4271      ^4281      ^4291      

TCAGTCACGC CTTTTCTAGG GCAACAGCAA CAGCTGCAAT CATTAGTGCC ACTGACCCCG TCTGCTTTGT TTGGTGGCGC CAATTTTAAT CAAAGTGGGA 
^4301      ^4311      ^4321      ^4331      ^4341      ^4351      ^4361      ^4371      ^4381      ^4391      

ATATTGCTGA TAGCTCATTG TCCTTCACTT TCACTAACAG TAGCAACGGT CCGAACCTCA TAACAACTCA AACAAATTCT CAAGCGCTTT CACAACCAAT 
^4401      ^4411      ^4421      ^4431      ^4441      ^4451      ^4461      ^4471      ^4481      ^4491      

TGCCTCCTCT AACGTTCATG ATAACTTCAT GAATAATGAA ATCACGGCTA GTAAAATTGA TGATGGTAAT AATTCAAAAC CACTGTCACC TGGTTGGACG 
^4501      ^4511      ^4521      ^4531      ^4541      ^4551      ^4561      ^4571      ^4581      ^4591      

GACCAAACTG CGTATAACGC GTTTGGAATC ACTACAGGGA TGTTTAATAC CACTACAATG GATGATGTAT ATAACTATCT ATTCGATGAT GAAGATACCC 
^4601      ^4611      ^4621      ^4631      ^4641      ^4651      ^4661      ^4671      ^4681      ^4691      

CACCAAACCC AAAAAAAGAG TAAGAGCTCA GCTAGCTAAC TGAATAAGGA ACAATGAACG TTTTTCCTTT CTCTTGTTCC TAGTATTAAT GACTGACCGA 
^4701      ^4711      ^4721      ^4731      ^4741      ^4751      ^4761      ^4771      ^4781      ^4791      

TACATCCCTT TTTTTTTTTG TCTTTGTCTA GCTCCAATTC GCCCTATAGT GAGTCGTATT ACAATTCACT GGCCGTCGTT TTACAACGTC GTGACTGGGA 
^4801      ^4811      ^4821      ^4831      ^4841      ^4851      ^4861      ^4871      ^4881      ^4891      

AAACCCTGGC GTTACCCAAC TTAATCGCCT TGCAGCACAT CCCCCTTTCG CCAGCTGGCG TAATAGCGAA GAGGCCCGCA CCGATCGCCC TTCCCAACAG 
^4901      ^4911      ^4921      ^4931      ^4941      ^4951      ^4961      ^4971      ^4981      ^4991      

TTGCGCAGCC TGAATGGCGA ATGGACGCGC CCTGTAGCGG CGCATTAAGC GCGGCGGGTG TGGTGGTTAC GCGCAGCGTG ACCGCTACAC TTGCCAGCGC 
^5001      ^5011      ^5021      ^5031      ^5041      ^5051      ^5061      ^5071      ^5081      ^5091      

CCTAGCGCCC GCTCCTTTCG CTTTCTTCCC TTCCTTTCTC GCCACGTTCG CCGGCTTTCC CCGTCAAGCT CTAAATCGGG GGCTCCCTTT AGGGTTCCGA 
^5101      ^5111      ^5121      ^5131      ^5141      ^5151      ^5161      ^5171      ^5181      ^5191      

TTTAGTGCTT TACGGCACCT CGACCCCAAA AAACTTGATT AGGGTGATGG TTCACGTAGT GGGCCATCGC CCTGATAGAC GGTTTTTCGC CCTTTGACGT 
^5201      ^5211      ^5221      ^5231      ^5241      ^5251      ^5261      ^5271      ^5281      ^5291      

TGGAGTCCAC GTTCTTTAAT AGTGGACTCT TGTTCCAAAC TGGAACAACA CTCAACCCTA TCTCGGTCTA TTCTTTTGAT TTATAAGGGA TTTTGCCGAT 
^5301      ^5311      ^5321      ^5331      ^5341      ^5351      ^5361      ^5371      ^5381      ^5391      

TTCGGCCTAT TGGTTAAAAA ATGAGCTGAT TTAACAAAAA TTTAACGCGA ATTTTAACAA AATATTAACG CTTACAATTT CCTGATGCGG TATTTTCTCC 
^5401      ^5411      ^5421      ^5431      ^5441      ^5451      ^5461      ^5471      ^5481      ^5491      

TTACGCATCT GTGCGGTATT TCACACCGCA TAGGGTAATA ACTGATATAA TTAAATTGAA GCTCTAATTT GTGAGTTTAG TATACATGCA TTTACTTATA 
^5501      ^5511      ^5521      ^5531      ^5541      ^5551      ^5561      ^5571      ^5581      ^5591      

ATACAGTTTT TTAGTTTTGC TGGCCGCATC TTCTCAAATA TGCTTCCCAG CCTGCTTTTC TGTAACGTTC ACCCTCTACC TTAGCATCCC TTCCCTTTGC 
^5601      ^5611      ^5621      ^5631      ^5641      ^5651      ^5661      ^5671      ^5681      ^5691      

AAATAGTCCT CTTCCAACAA TAATAATGTC AGATCCTGTA GAGACCACAT CATCCACGGT TCTATACTGT TGACCCAATG CGTCTCCCTT GTCATCTAAA 
^5701      ^5711      ^5721      ^5731      ^5741      ^5751      ^5761      ^5771      ^5781      ^5791      

CCCACACCGG GTGTCATAAT CAACCAATCG TAACCTTCAT CTCTTCCACC CATGTCTCTT TGAGCAATAA AGCCGATAAC AAAATCTTTG TCGCTCTTCG 
^5801      ^5811      ^5821      ^5831      ^5841      ^5851      ^5861      ^5871      ^5881      ^5891      

CAATGTCAAC AGTACCCTTA GTATATTCTC CAGTAGATAG GGAGCCCTTG CATGACAATT CTGCTAACAT CAAAAGGCCT CTAGGTTCCT TTGTTACTTC 
^5901      ^5911      ^5921      ^5931      ^5941      ^5951      ^5961      ^5971      ^5981      ^5991      

TTCTGCCGCC TGCTTCAAAC CGCTAACAAT ACCTGGCCCC ACCACACCGT GTGCATTCGT AATGTCTGCC CATTCTGCTA TTCTGTATAC ACCCGCAGAG 
^6001      ^6011      ^6021      ^6031      ^6041      ^6051      ^6061      ^6071      ^6081      ^6091      

TACTGCAATT TGACTGTATT ACCAATGTCA GCAAATTTTC TGTCTTCGAA GAGTAAAAAA TTGTACTTGG CGGATAATGC CTTTAGCGGC TTAACTGTGC 
^6101      ^6111      ^6121      ^6131      ^6141      ^6151      ^6161      ^6171      ^6181      ^6191      

CCTCCATCGA AAAATCAGTC AATATATCCA CATGTGTTTT TAGTAAACAA ATTTTGGGAC CTAATGCTTC AACTAACTCC AGTAATTCCT TGGTGGTACG 
^6201      ^6211      ^6221      ^6231      ^6241      ^6251      ^6261      ^6271      ^6281      ^6291      

AACATCCAAT GAAGCACACA AGTTTGTTTG CTTTTCGTGC ATGATATTAA ATAGCTTGGC AGCAACAGGA CTAGGATGAG TAGCAGCACG TTCCTTATAT 
^6301      ^6311      ^6321      ^6331      ^6341      ^6351      ^6361      ^6371      ^6381      ^6391      

GTAGCTTTCG ACATGATTTA TCTTCGTTTC CTGCAGGTTT TTGTTCTGTG CAGTTGGGTT AAGAATACTG GGCAATTTCA TGTTTCTTCA ACACTACATA 
^6401      ^6411      ^6421      ^6431      ^6441      ^6451      ^6461      ^6471      ^6481      ^6491      

TGCGTATATA TACCAATCTA AGTCTGTGCT CCTTCCTTCG TTCTTCCTTC TGTTCGGAGA TTACCGAATC AAAAAAATTT CAAGGAAACC GAAATCAAAA 
^6501      ^6511      ^6521      ^6531      ^6541      ^6551      ^6561      ^6571      ^6581      ^6591      

AAAAGAATAA AAAAAAAATG ATGAATTGAA AAGGTGGTAT GGTGCACTCT CAGTACAATC TGCTCTGATG CCGCATAGTT AAGCCAGCCC CGACACCCGC 
^6601      ^6611      ^6621      ^6631      ^6641      ^6651      ^6661      ^6671      ^6681      ^6691      

CAACACCCGC TGACGCGCCC TGACGGGCTT GTCTGCTCCC GGCATCCGCT TACAGACAAG CTGTGACCGT CTCCGGGAGC TGCATGTGTC AGAGGTTTTC 
^6701      ^6711      ^6721      ^6731      ^6741      ^6751      ^6761      ^6771      ^6781      ^6791      

ACCGTCATCA CCGAAACGCG CGAGACGAAA GGGCCTCGTG ATACGCCTAT TTTTATAGGT TAATGTCATG ATAATAATGG TTTCTTAGAC GTGCGGCCGC 
^6801      ^6811      ^6821      ^6831      ^6841      ^6851      ^6861      ^6871      ^6881      ^6891      

TCTAGAACTA GTGGATCAAT TCCACGGACT ATAGACTATA CTAGTATACT CCGTCTACTG TACGATACAC TTCCGCTCAG GTCCTTGTCC TTTAACGAGG 
^6901      ^6911      ^6921      ^6931      ^6941      ^6951      ^6961      ^6971      ^6981      ^6991      

CCTTACCACT CTTTTGTTAC TCTATTGATC CAGCTCAGCA AAGGCAGTGT GATCTAAGAT TCTATCTTCG CGATGTAGTA AAACTAGCTA GACCGAGAAA 
^7001      ^7011      ^7021      ^7031      ^7041      ^7051      ^7061      ^7071      ^7081      ^7091      

GAGACTAGAA ATGCAAAAGG CACTTCTACA ATGGCTGCCA TCATTATTAT CCGATGTGAC GCTGCAGCTT CTCAATGATA TTCGAATACG CTTTGAGGAG 
^7101      ^7111      ^7121      ^7131      ^7141      ^7151      ^7161      ^7171      ^7181      ^7191      

ATACAGCCTA ATATCCGACA AACTGTTTTA CAGATTTACG ATCGTACTTG TTACCCATCA TTGAATTTTG AACATCCGAA CCTGGGAGTT TTCCCTGAAA 
^7201      ^7211      ^7221      ^7231      ^7241      ^7251      ^7261      ^7271      ^7281      ^7291      

CAGATAGTAT ATTTGAACCT GTATAATAAT ATATAGTCTA GCGCTTTACG GAAGACAATG TATGTATTTC GGTTCCTGGA GAAACTATTG CATCTATTGC 
^7301      ^7311      ^7321      ^7331      ^7341      ^7351      ^7361      ^7371      ^7381      ^7391      

ATAGGTAATC TTGCACGTCG CATCCCCGGT TCATTTTCTG CGTTTCCATC TTGCACTTCA ATAGCATATC TTTGTTAACG AAGCATCTGT GCTTCATTTT 
^7401      ^7411      ^7421      ^7431      ^7441      ^7451      ^7461      ^7471      ^7481      ^7491      

GTAGAACAAA AATGCAACGC GAGAGCGCTA ATTTTTCAAA CAAAGAATCT GAGCTGCATT TTTACAGAAC AGAAATGCAA CGCGAAAGCG CTATTTTACC 
^7501      ^7511      ^7521      ^7531      ^7541      ^7551      ^7561      ^7571      ^7581      ^7591      

AACGAAGAAT CTGTGCTTCA TTTTTGTAAA ACAAAAATGC AACGCGAGAG CGCTAATTTT TCAAACAAAG AATCTGAGCT GCATTTTTAC AGAACAGAAA 
^7601      ^7611      ^7621      ^7631      ^7641      ^7651      ^7661      ^7671      ^7681      ^7691      

TGCAACGCGA GAGCGCTATT TTACCAACAA AGAATCTATA CTTCTTTTTT GTTCTACAAA AATGCATCCC GAGAGCGCTA TTTTTCTAAC AAAGCATCTT 
^7701      ^7711      ^7721      ^7731      ^7741      ^7751      ^7761      ^7771      ^7781      ^7791      

AGATTACTTT TTTTCTCCTT TGTGCGCTCT ATAATGCAGT CTCTTGATAA CTTTTTGCAC TGTAGGTCCG TTAAGGTTAG AAGAAGGCTA CTTTGGTGTC 
^7801      ^7811      ^7821      ^7831      ^7841      ^7851      ^7861      ^7871      ^7881      ^7891      

TATTTTCTCT TCCATAAAAA AAGCCTGACT CCACTTCCCG CGTTTACTGA TTACTAGCGA AGCTGCGGGT GCATTTTTTC AAGATAAAGG CATCCCCGAT 
^7901      ^7911      ^7921      ^7931      ^7941      ^7951      ^7961      ^7971      ^7981      ^7991      

TATATTCTAT ACCGATGTGG ATTGCGCATA CTTTGTGAAC AGAAAGTGAT AGCGTTGATG ATTCTTCATT GGTCAGAAAA TTATGAACGG TTTCTTCTAT 
^8001      ^8011      ^8021      ^8031      ^8041      ^8051      ^8061      ^8071      ^8081      ^8091      

TTTGTCTCTA TATACTACGT ATAGGAAATG TTTACATTTT CGTATTGTTT TCGATTCACT CTATGAATAG TTCTTACTAC AATTTTTTTG TCTAAAGAGT 
^8101      ^8111      ^8121      ^8131      ^8141      ^8151      ^8161      ^8171      ^8181      ^8191      

AATACTAGAG ATAAACATAA AAAATGTAGA GGTCGAGTTT AGATGCAAGT TCAAGGAGCG AAAGGTGGAT GGGTAGGTTA TATAGGGATA TAGCACAGAG 
^8201      ^8211      ^8221      ^8231      ^8241      ^8251      ^8261      ^8271      ^8281      ^8291      

ATATATAGCA AAGAGATACT TTTGAGCAAT GTTTGTGGAA GCGGTATTCG CAATATTTTA GTAGCTCGTT ACAGTCCGGT GCGTTTTTGG TTTTTTGAAA 
^8301      ^8311      ^8321      ^8331      ^8341      ^8351      ^8361      ^8371      ^8381      ^8391      

GTGCGTCTTC AGAGCGCTTT TGGTTTTCAA AAGCGCTCTG AAGTTCCTAT ACTTTCTAGA GAATAGGAAC TTCGGAATAG GAACTTCAAA GCGTTTCCGA 
^8401      ^8411      ^8421      ^8431      ^8441      ^8451      ^8461      ^8471      ^8481      ^8491      

AAACGAGCGC TTCCGAAAAT GCAACGCGAG CTGCGCACAT ACAGCTCACT GTTCACGTCG CACCTATATC TGCGTGTTGC CTGTATATAT ATATACATGA 
^8501      ^8511      ^8521      ^8531      ^8541      ^8551      ^8561      ^8571      ^8581      ^8591      

GAAGAACGGC ATAGTGCGTG TTTATGCTTA AATGCGTACT TATATGCGTC TATTTATGTA GGATGAAAGG TAGTCTAGTA CCTCCTGTGA TATTATCCCA 
^8601      ^8611      ^8621      ^8631      ^8641      ^8651      ^8661      ^8671      ^8681      ^8691      

TTCCATGCGG GGTATCGTAT GCTTCCTTCA GCACTACCCT TTAGCTGTTC TATATGCTGC CACTCCTCAA TTGGATTAGT CTCATCCTTC AATGCTATCA 
^8701      ^8711      ^8721      ^8731      ^8741      ^8751      ^8761      ^8771      ^8781      ^8791      

TTTCCTTTGA TATTGGATCA TATGCATAGT ACCGAGAAAC TAGTGCGAAG TAGTGATCAG GTATTGCTGT TATCTGATGA GTATACGTTG TCCTGGCCAC 
^8801      ^8811      ^8821      ^8831      ^8841      ^8851      ^8861      ^8871      ^8881      ^8891      

GGCAGAAGCA CGCTTATCGC TCCAATTTCC CACAACATTA GTCAACTCCG TTAGGCCCTT CATTGAAAGA AATGAGGTCA TCAAATGTCT TCCAATGTGA 
^8901      ^8911      ^8921      ^8931      ^8941      ^8951      ^8961      ^8971      ^8981      ^8991      

GATTTTGGGC CATTTTTTAT AGCAAAGATT GAATAAGGCG CATTTTTCTT CAAAGCTGCG GCCGCACTCT CACTAGTACG TCAGGTGGCA CTTTTCGGGG 
^9001      ^9011      ^9021      ^9031      ^9041      ^9051      ^9061      ^9071      ^9081      ^9091      

AAATGTGCGC GGAACCCCTA TTTGTTTATT TTTCTAAATA CATTCAAATA TGTATCCGCT CATGAGACAA TAACCCTGAT AAATGCTTCA ATAATATTGA 
^9101      ^9111      ^9121      ^9131      ^9141      ^9151      ^9161      ^9171      ^9181      ^9191      

AAAAGGAAGA GTATGAGTAT TCAACATTTC CGTGTCGCCC TTATTCCCTT TTTTGCGGCA TTTTGCCTTC CTGTTTTTGC TCACCCAGAA ACGCTGGTGA 
^9201      ^9211      ^9221      ^9231      ^9241      ^9251      ^9261      ^9271      ^9281      ^9291      

AAGTAAAAGA TGCTGAAGAT CAGTTGGGTG CACGAGTGGG TTACATCGAA CTGGATCTCA ACAGCGGTAA GATCCTTGAG AGTTTTCGCC CCGAAGAACG 
^9301      ^9311      ^9321      ^9331      ^9341      ^9351      ^9361      ^9371      ^9381      ^9391      

TTTTCCAATG ATGAGCACTT TTAAAGTTCT GCTATGTGGC GCGGTATTAT CCCGTATTGA CGCCGGGCAA GAGCAACTCG GTCGCCGCAT ACACTATTCT 
^9401      ^9411      ^9421      ^9431      ^9441      ^9451      ^9461      ^9471      ^9481      ^9491      

CAGAATGACT TGGTTGAGTA CTCACCAGTC ACAGAAAAGC ATCTTACGGA TGGCATGACA GTAAGAGAAT TATGCAGTGC TGCCATAACC ATGAGTGATA 
^9501      ^9511      ^9521      ^9531      ^9541      ^9551      ^9561      ^9571      ^9581      ^9591      

ACACTGCGGC CAACTTACTT CTGACAACGA TCGGAGGACC GAAGGAGCTA ACCGCTTTTT TGCACAACAT GGGGGATCAT GTAACTCGCC TTGATCGTTG 
^9601      ^9611      ^9621      ^9631      ^9641      ^9651      ^9661      ^9671      ^9681      ^9691      

GGAACCGGAG CTGAATGAAG CCATACCAAA CGACGAGCGT GACACCACGA TGCCTGTAGC AATGGCAACA ACGTTGCGCA AACTATTAAC TGGCGAACTA 
^9701      ^9711      ^9721      ^9731      ^9741      ^9751      ^9761      ^9771      ^9781      ^9791      

CTTACTCTAG CTTCCCGGCA ACAATTAATA GACTGGATGG AGGCGGATAA AGTTGCAGGA CCACTTCTGC GCTCGGCCCT TCCGGCTGGC TGGTTTATTG 
^9801      ^9811      ^9821      ^9831      ^9841      ^9851      ^9861      ^9871      ^9881      ^9891      

CTGATAAATC TGGAGCCGGT GAGCGTGGGT CTCGCGGTAT CATTGCAGCA CTGGGGCCAG ATGGTAAGCC CTCCCGTATC GTAGTTATCT ACACGACGGG 
^9901      ^9911      ^9921      ^9931      ^9941      ^9951      ^9961      ^9971      ^9981      ^9991      

GAGTCAGGCA ACTATGGATG AACGAAATAG ACAGATCGCT GAGATAGGTG CCTCACTGAT TAAGCATTGG TAACTGTCAG ACCAAGTTTA CTCATATATA 
^10001     ^10011     ^10021     ^10031     ^10041     ^10051     ^10061     ^10071     ^10081     ^10091     

CTTTAGATTG ATTTAAAACT TCATTTTTAA TTTAAAAGGA TCTAGGTGAA GATCCTTTTT GATAATCTCA TGACCAAAAT CCCTTAACGT GAGTTTTCGT 
^10101     ^10111     ^10121     ^10131     ^10141     ^10151     ^10161     ^10171     ^10181     ^10191     

TCCACTGAGC GTCAGACCCC GTAGAAAAGA TCAAAGGATC TTCTTGAGAT CCTTTTTTTC TGCGCGTAAT CTGCTGCTTG CAAACAAAAA AACCACCGCT 
^10201     ^10211     ^10221     ^10231     ^10241     ^10251     ^10261     ^10271     ^10281     ^10291     

ACCAGCGGTG GTTTGTTTGC CGGATCAAGA GCTACCAACT CTTTTTCCGA AGGTAACTGG CTTCAGCAGA GCGCAGATAC CAAATACTGT TCTTCTAGTG 
^10301     ^10311     ^10321     ^10331     ^10341     ^10351     ^10361     ^10371     ^10381     ^10391     

TAGCCGTAGT TAGGCCACCA CTTCAAGAAC TCTGTAGCAC CGCCTACATA CCTCGCTCTG CTAATCCTGT TACCAGTGGC TGCTGCCAGT GGCGATAAGT 
^10401     ^10411     ^10421     ^10431     ^10441     ^10451     ^10461     ^10471     ^10481     ^10491     

CGTGTCTTAC CGGGTTGGAC TCAAGACGAT AGTTACCGGA TAAGGCGCAG CGGTCGGGCT GAACGGGGGG TTCGTGCACA CAGCCCAGCT TGGAGCGAAC 
^10501     ^10511     ^10521     ^10531     ^10541     ^10551     ^10561     ^10571     ^10581     ^10591     

GACCTACACC GAACTGAGAT ACCTACAGCG TGAGCTATGA GAAAGCGCCA CGCTTCCCGA AGGGAGAAAG GCGGACAGGT ATCCGGTAAG CGGCAGGGTC 
^10601     ^10611     ^10621     ^10631     ^10641     ^10651     ^10661     ^10671     ^10681     ^10691     

GGAACAGGAG AGCGCACGAG GGAGCTTCCA GGGGGAAACG CCTGGTATCT TTATAGTCCT GTCGGGTTTC GCCACCTCTG ACTTGAGCGT CGATTTTTGT 
^10701     ^10711     ^10721     ^10731     ^10741     ^10751     ^10761     ^10771     ^10781     ^10791     

GATGCTCGTC AGGGGGGCGG AGCCTATGGA AAAACGCCAG CAACGCGGCC TTTTTACGGT TCCTGGCCTT TTGCTGGCCT TTTGCTCACA TGTTCTTTCC 
^10801     ^10811     ^10821     ^10831     ^10841     ^10851     ^10861     ^10871     ^10881     ^10891     

TGCGTTATCC CCTGATTCTG TGGATAACCG TATTACCGCC TTTGAGTGAG CTGATACCGC TCGCCGCAGC CGAACGACCG AGCGCAGCGA GTCAGTGAGC 
^10901     ^10911     ^10921     ^10931     ^10941     ^10951     ^10961     ^10971     ^10981     ^10991     

GAGGAAGCGG AAGAGCGCCC AATACGCAAA CCGCCTCTCC CCGCGCGTTG GCCGATTCAT TAATGCAGCT GGCACGACAG GTTTCCCGAC TGGAAAGCGG 
^11001     ^11011     ^11021     ^11031     ^11041     ^11051     ^11061     ^11071     ^11081     ^11091     

GCAGTGAGCG CAACGCAATT AATGTGAGTT AGCTCACTCA TTAGGCACCC CAGGCTTTAC ACTTTATGCT TCCGGCTCGT ATGTTGTGTG GAATTGTGAG 
^11101     ^11111     ^11121     ^11131     ^11141     ^11151     ^11161     ^11171     ^11181     ^11191     

CGGATAACAA TTTCACACAG GAAACAGCTA TGACCATGAT TACGCCAAGC TCGAAATTAA CCCTCACTAA AGGGAACAAA AGCTGGTACC GGGCCGGCCG 
^11201     ^11211     ^11221     ^11231     ^11241     ^11251     ^11261     ^11271     ^11281     ^11291     

TCGAGCTTGA TGGCATCGTG GTGTCACGCT CGTCGTTTGG TATGGCTTCA TTCAGCTCCG GTTCCCAACG ATCAAGGCGA GTTACATGAT CCCCCATGTT 
^11301     ^11311     ^11321     ^11331     ^11341     ^11351     ^11361     ^11371     ^11381     ^11391     

GTGCAAAAAA GCGGTTAGCT CCTTCGGTCC TCCGATCGTT GTCAGAAGTA AGTTGGCCGC AGTGTTATCA CTCATGGTTA TGGCAGCACT GCATAATTCT 
^11401     ^11411     ^11421     ^11431     ^11441     ^11451     ^11461     ^11471     ^11481     ^11491     

CTTACTGTCA TGCCATCCGT AAGATGCTTT TCTGTGACTG GTGAGTACTC AACCAAGTCA TTCTGAGAAT AGTGTATGCG GCGACCGAGT TGCTCTTGCC 
^11501     ^11511     ^11521     ^11531     ^11541     ^11551     ^11561     ^11571     ^11581     ^11591     

CGGCGTCAAC ACGGGATAAT ACCGCGCCAC ATAGCAGAAC TTTAAAAGTG CTCATCATTG GAAAACGTTC TTCGGGGCGA AAACTCTCAA GGATCTTACC 
^11601     ^11611     ^11621     ^11631     ^11641     ^11651     ^11661     ^11671     ^11681     ^11691     

GCTGTTGAGA TCCAGTTCGA TGTAACCCAC TCGTGCACCC AACTGATCTT CAGCATCTTT TACTTTCACC AGCGTTTCTG GGTGAGCAAA AACAGGAAGG 
^11701     ^11711     ^11721     ^11731     ^11741     ^11751     ^11761     ^11771     ^11781     ^11791     

CAAAATGCCG CAAAAAAGGG AATAAGGGCG ACACGGAAAT GTTGAATACT CATACTCTTC CTTTTTCAAT ATTATTGAAG CATTTATCAG GGTTATTGTC 
^11801     ^11811     ^11821     ^11831     ^11841     ^11851     ^11861     ^11871     ^11881     ^11891     

TCATGAGCGG ATACATATTT GAATGTATTT AGAAAAATAA ACAAATAGGG GTTCCGCGCA CATTTCCCCG AAAAGTGCCA CCTGACGTCT AAGAAACCAT 
^11901     ^11911     ^11921     ^11931     ^11941     ^11951     ^11961     ^11971     ^11981     ^11991     

TATTATCATG ACATTAACCT ATAAAAATAG GCGTATCACG AGGCCCTTTC GTCTTCAAGA ATTGGGGATC TACGTATGGT CATTTCTTCT TCAGATTCCC 
^12001     ^12011     ^12021     ^12031     ^12041     ^12051     ^12061     ^12071     ^12081     ^12091     

TCATGGAGAA AGTGCGGCAG ATGTATATGA CAGAGTCGCC AGTTTCCAAG AGACTTTATT CAGGCACTTC CATGATAGGC AAGAGAGAAG ACCCAGAGAT 
^12101     ^12111     ^12121     ^12131     ^12141     ^12151     ^12161     ^12171     ^12181     ^12191     

GTTGTTGTCC TAGTTACACA TGGTATTTAT TCCAGAGTAT TCCTGATGAA ATGGTTTAGA TGGACATACG AAGAGTTTGA ATCGTTTACC AATGTTCCTA 
^12201     ^12211     ^12221     ^12231     ^12241     ^12251     ^12261     ^12271     ^12281     ^12291     

ACGGGAGCGT AATGGTGATG GAACTGGACG AATCCATCAA TAGATACGTC CTGAGGACCG TGCTACCCAA ATGGACTGAT TGTGAGGGAG ACCTAACTAC 
^12301     ^12311     ^12321     ^12331     ^12341     ^12351     ^12361     ^12371     ^12381     ^12391     

ATAGTGTTTA AAGATTACGG ATATTTAACT TACTTAGAAT AATGCCATTT TTTTGAGTTA TAATAATCCT ACGTTAGTGT GAGCGGGATT TAAACTGTGA 
^12401     ^12411     ^12421     ^12431     ^12441     ^12451     ^12461     ^12471     ^12481     ^12491     

GGACCTTAAT ACATTCAGAC ACTTCTGCGG TATCACCCTA CTTATTCCCT TCGAGATTAT ATCTAGGAAC CCATCAGGTT GGTGGAAGAT TACCCGTTCT 
^12501     ^12511     ^12521     ^12531     ^12541     ^12551     ^12561     ^12571     ^12581     ^12591     

AAGACTTTTC AGCTTCCTCT ATTGATGTTA CACCTGGACA CCCCTTTTCT GGCATCCAGT TTTTAATCTT CAGTGGCATG TGAGATTCTC CGAAATTAAT 
^12601     ^12611     ^12621     ^12631     ^12641     ^12651     ^12661     ^12671     ^12681     ^12691     

TAAAGCAATC ACACAATTCT CTCGGATACC ACCTCGGTTG AAACTGACAG GTGGTTTGTT ACGCATGCTA ATGCAAAGGA GCCTATATAC CTTTGGCTCG 
^12701     ^12711     ^12721     ^12731     ^12741     ^12751     ^12761     ^12771     ^12781     ^12791     

GCTGCTGTAA CAGGGAATAT AAAGGGCAGC ATAATTTAGG AGTTTAGTGA ACTTGCAACA TTTACTATTT TCCCTTCTTA CGTAAATATT TTTCTTTTTA 
^12801     ^12811     ^12821     ^12831     ^12841     ^12851     ^12861     ^12871     ^12881     ^12891     

ATTCTAAATC AATCTTTTTC AATTTTTTGT TTGTATTCTT TTCTTGCTTA AATCTATAAC TACAAAAAAC ACATACAG
^12901     ^12911     ^12921     ^12931     ^12941     ^12951     ^12961     ^12971

Plasmid sequence verified (partially/completely) by IRESite (more details in Remarks):

plasmid sequence confirmed by IRESite curators by restriction analysis + parts by PCR + sequencing

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: Tomáš Mašek	Reviewed by: Tomáš Mašek

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:

no

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

1-1653

ORF

ORF position:

2

Version: `1`
Originaly submitted by: Tomáš Mašek	Reviewed by: Tomáš Mašek

The abbreviated name of this ORF/gene:
HCV1-GAL4

The description of the protein encoded in this ORF:
The fusion protein comprises the first 15 amino acids of HCV viral polyprotein and GAL4 DNA-binding transcription factor. Gal4p is required for the activation of the GAL genes in response to galactose; repressed by Gal80p and activated by Gal3p.

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:

yes

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

2065-4710

OPTIONAL: The function of the encoded protein

Molecular Function: transcriptional activator activity (TAS)
Biological Process: galactose metabolism (TAS) regulation of transcription, DNA-dependent (TAS)
Cellular Component: nucleus (TAS)

Remarks:

Sc_HCV_type_1a IRES was inserted into intercistronic position of pFGAL4 bicistronic plasmid that can be used
in the pFGAL4h/PJ69A reporter system. pFGAL4 vector bears genes for the firefly luciferase and Gal4p as a
first and second cistron, respectively. The yeast strain PJ69A contains besides the GAL4 and GAL80 deletions
also reporter genes coding for the yeast Ade2 and His3 proteins and for the bacterial beta-galactosidase - all
of them under the control of the Gal4-inducible Gal1 promoter. Thus pFGAL4/PJ69A represents a specialised and
sensitive system, which allows an enhancement of the measured signal by in vivo coupled transcription and
enzymatic detection. This system can be used for both measuring the ratio of the beta-gal/luc enzymatic
activities and searching for IRES activity by screening the colony growth rates on selection media lacking
adenine and histidine and containing various concentrations of the competitive inhibitor of the histidine
biosynthetic pathway.

Vector was created by IRESite curator - thus whole sequence is verified by above mentioned methods.

Citations:

Masek T., Vopalensky V., Horvath O., Vortelova L., Feketova Z., Pospisek M. (2007) Hepatitis C virus internal ribosome entry site initiates protein synthesis at the authentic initiation codon in yeast. J. Gen. Virol. 88(Pt 7):1992-2002

IRESs:

IRES:

Version: `3`	Last change: `2008-07-03 19:30:42`
Originaly submitted by: Tomáš Mašek	Reviewed by: Tomáš Mašek
The IRES name: `HCV_type_1a`	The functional status of IRES: `functional`

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

1670-2054

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: `401`
5'-end of IRES relative to first base of the START codon of the downstream ORF: `-395`	3'-end of IRES relative to first base of the START codon of the downstream ORF: `-11`

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

GCCAGCCCCC TGATGGGGGC GACACTCCAC CATGAATCAC TCCCCTGTGA GGAACTACTG TCTTCACGCA GAAAGCGTCT AGCCATGGCG TTAGTATGAG 
^1670      ^1680      ^1690      ^1700      ^1710      ^1720      ^1730      ^1740      ^1750      ^1760      

TGTCGTGCAG CCTCCAGGAC CCCCCCTCCC GGGAGAGCCA TAGTGGTCTG CGGAACCGGT GAGTACACCG GAATTGCCAG GACGACCGGG TCCTTTCTTG 
^1770      ^1780      ^1790      ^1800      ^1810      ^1820      ^1830      ^1840      ^1850      ^1860      

GATAAACCCG CTCAATGCCT GGAGATTTGG GCGTGCCCCC GCAAGACTGC TAGCCGAGTA GTGTTGGGTC GCGAAAGGCC TTGTGGTACT GCCTGATAGG 
^1870      ^1880      ^1890      ^1900      ^1910      ^1920      ^1930      ^1940      ^1950      ^1960      

GTGCTTGCGA GTGCCCCGGG AGGTCTCGTA GACCGTGCAC CATGAGCACG AATCCTAAAC CTCAAAGAAA AACCAAACGT AACAC
^1970      ^1980      ^1990      ^2000      ^2010      ^2020      ^2030      ^2040      ^2050

Remarks:

Sc_HCV_type_1a IRES represents the full-length 5'-UTR of HCV RNA (type 1a) and the first 15 codons of viral
polyprotein.

Citations:

The translation experiments:

Translation results:

IRESite Translation Id: 81

Version: `4`	Last change: `2008-07-03 19:30:42`
Originaly submitted by: Tomáš Mašek	Reviewed by: Tomáš Mašek

The translation method used to study IRES function:
in vivo

The organism used for translation:

Saccharomyces cerevisiae pJ69a

The temperature (in degrees of Celsia):
28

The relative translation efficiency in % of this IRES:

92.000

Name of IRES used as the positive control:

HCV1a

Name of the plasmid used as the positive control. `pFGAL4h-HCV1`	Name of the plasmid used as the negative control. `pFGAL4-L428`
`IRESite Id` of the plasmid used as positive control. `86`	`IRESite Id` of the plasmid used as negative control. `126`
The relative translation efficiency in % of the positive control: `100.000`	The relative translation efficiency in % of the negative control: `3.800`
The size (length) of intercistronic region in the positive control: `453`	The size (length) of intercistronic region in the negative control: `432`

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:

We compared Gal4p translation efficiency in yeast clones bearing either pFGAL4h-HCV1 or pFGAL4-HCV1 vectors,
containing and not containing the hairpin loop respectively and confirmed that the presence of the stable
hairpin loop preceding HCV IRES has almost no influence on its activity. The relative translation efficiency
and size of intercistronic region of negative control have been inserted as an avarage values of all negative
controls used (pFGAL4-L413,L-428, L456, IDs 126-128).

Citations:

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