Supporting Information Sitaraman et al. 10.1073/pnas.1102883108
WTREP dREP sREP
Fig. S1.
•321 GGTTTGAACGCGCAGCCGCCATGCCGGGGTTTTACGAGATTGTGATTAAGGTCCCCAGCG 360 --------------------ATGCCCGGGTTTTACGAGATCGTGATTAAGGTGCCATCCG --------------------ATGCCCGGATTCTACGAAATCGTCATCAAAGTGCCCTCTG ***** ** ** ***** ** ** ** ** ** ** *
WTREP dREP sREP
ACCTTGACGAGCATCTGCCCGGCATTTCTGACAGCTTTGTGAACTGGGTGGCCGAGAAGG 420 ATCTCGACGAGCATCTGCCCGGGATTAGCGATTCGTTCGTGAATTGGGTCGCCGAAAAGG ACTTGGATGAACACCTGCCGGGGATCAGCGATTCTTTCGTCAATTGGGTCGCGGAGAAAG * * ** ** ** ***** ** ** ** ** ** ** ***** ** ** ** *
WTREP dREP sREP
AATGGGAGTTGCCGCCAGATTCTGACATGGATCTGAATCTGATTGAGCAGGCACCCCTGA 480 AGTGGGAGTTGCCCCCCGATAGCGATATGGACCTGAATCTGATCGAGCAGGCCCCCCTTA AGTGGGAACTTCCCCCCGACTCGGACATGGACCTGAACTTAATCGAGCAAGCCCCGCTGA * ***** * ** ** ** ** ***** ***** * ** ***** ** ** ** *
WTREP dREP sREP
CCGTGGCCGAGAAGCTGCAGCGCGACTTTCTGACGGAATGGCGCCGTGTGAGTAAGGCCC 540 CCGTCGCCGAGAAACTGCAACGCGATTTCTTGACCGAGTGGAGACGCGTGAGTAAGGCCC CGGTGGCGGAGAAACTGCAGCGGGACTTTCTGACCGAGTGGAGGCGCGTATCGAAAGCGC * ** ** ***** ***** ** ** ** **** ** *** * ** ** ** ** *
WTREP dREP sREP
CGGAGGCCCTTTTCTTTGTGCAATTTGAGAAGGGAGAGAGCTACTTCCACATGCACGTGC 600 CCGAAGCCCTGTTTTTCGTGCAATTTGAAAAGGGCGAGTCATACTTTCATATGCACGTGT CCGAAGCTTTGTTTTTCGTCCAGTTCGAGAAGGGGGAGTCGTACTTTCATATGCATGTGT * ** ** * ** ** ** ** ** ** ***** *** ***** ** ***** ***
WTREP dREP sREP
TCGTGGAAACCACCGGGGTGAAATCCATGGTTTTGGGACGTTTCCTGAGTCAGATTCGCG 660 TGGTCGAGACTACCGGCGTTAAGTCTATGGTGCTCGGACGGTTTCTGTCACAGATACGCG TGGTGGAGACTACGGGAGTGAAGAGTATGGTGCTAGGGAGGTTTCTGTCGCAAATAAGAG * ** ** ** ** ** ** ** ***** * ** * ** *** ** ** * *
WTREP dREP sREP
AAAAACTGATTCAGAGAATTTACCGCGGGATCGAGCCGACTTTGCCAAACTGGTTCGCGG 720 AAAAACTGATCCAGCGTATCTATCGCGGAATCGAGCCAACCCTACCGAATTGGTTCGCCG AGAAGCTGATCCAGCGGATATACCGTGGCATTGAGCCCACCCTTCCCAATTGGTTTGCCG * ** ***** *** * ** ** ** ** ** ***** ** * ** ** ***** ** *
WTREP dREP sREP
TCACAAAGACCAGAAATGGCGCCGGAGGCGGGAACAAGGTGGTGGATGAGTGCTACATCC 780 TTACGAAGACCCGTAACGGCGCCGGGGGGGGGAATAAGGTGGTCGACGAGTGCTATATCC TGACCAAAACTCGTAACGGAGCAGGGGGGGGAAATAAAGTCGTCGACGAGTGCTATATTC * ** ** ** * ** ** ** ** ** ** ** ** ** ** ** ******** ** *
WTREP dREP sREP
CCAATTACTTGCTCCCCAAAACCCAGCCTGAGCTCCAGTGGGCGTGGACTAATATGGAAC 840 CTAACTATCTGTTACCGAAAACGCAACCCGAGTTGCAGTGGGCCTGGACTAACATGGAGC CGAACTACCTCTTGCCCAAGACGCAGCCCGAATTGCAGTGGGCCTGGACCAACATGGAGC * ** ** * * ** ** ** ** ** ** * ******** ***** ** ***** *
WTREP dREP sREP
AGTATTTAAGCGCCTGTTTGAATCTCACGGAGCGTAAACGGTTGGTGGCGCAGCATCTGA 900 AATACTTGTCCGCATGCCTGAATCTGACCGAACGCAAACGGTTGGTCGCCCAGCATCTGA AATACCTGTCAGCGTGCCTCAACTTGACCGAAAGAAAGAGACTCGTGGCCCAGCACCTGA * ** * ** ** * ** * ** ** * ** * * ** ** ***** ****
WTREP dREP sREP
CGCACGTGTCGCAGACGCAGGAGCAGAACAAAGAGAATCAGAATCCCAATTCTGATGCGC 960 CACACGTGAGTCAGACCCAGGAGCAGAATAAGGAGAATCAGAATCCGAACTCCGACGCCC CCCATGTCTCACAGACCCAGGAACAGAATAAGGAAAACCAAAACCCAAATAGCGACGCCC * ** ** ***** ***** ***** ** ** ** ** ** ** ** ** ** *
(Continued)
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Fig. S1.
WTREP dREP sREP
CGGTGATCAGATCAAAAACTTCAGCCAGGTACATGGAGCTGGTCGGGTGGCTCGTGGACA 1020 CCGTGATACGGTCTAAGACTAGCGCTAGGTATATGGAGTTGGTGGGGTGGTTGGTCGACA CCGTGATACGGAGCAAGACCAGCGCTCGCTACATGGAGTTAGTGGGATGGTTGGTGGATA * ***** * ** ** ** * ** ****** * ** ** *** * ** ** *
WTREP dREP sREP
AGGGGATTACCTCGGAGAAGCAGTGGATCCAGGAGGACCAGGCCTCATACATCTCCTTCA 1080 AGGGGATTACCTCCGAGAAACAGTGGATCCAGGAGGACCAGGCGTCATACATTTCGTTTA AAGGAATCACGTCTGAGAAACAATGGATTCAGGAGGACCAGGCGTCCTACATTAGTTTTA * ** ** ** ** ***** ** ***** ************** ** ***** ** *
WTREP dREP sREP
ATGCGGCCTCCAACTCGCGG-TCCCAAATCAAGGCTGCCTTGGACAATGCGGGAAAGATT 1139 ACGCCGCATCGAACTCACGG-TCACAGATTAAGGCCGCACTCGACAACGCCGGTAAGATT ACGCCGCGTCAAA-TAGCAGATCTCAGATTAAAGCCGCGCTCGATAACGCCGGCAAAATC * ** ** ** ** * * * ** ** ** ** ** ** * ** ** ** ** ** **
WTREP dREP sREP
ATGAGCCTGACTAAAACCGCCCCCGACTACCTGGTGGGCCAGCAGCCCGTGGAGGACATT 1199 ATGAGTCTGACTAAGACCGCCCCCGATTACTTAGTGGGACAGCAACCCGTCGAGGACATT ATGTCGCTGACCAAGACAGCTCCCGACTACCTGGTGGGACAGCAGCCGGTGGAGGACATC *** ***** ** ** ** ***** *** * ***** ***** ** ** ********
WTREP dREP sREP
TCCAGCAATCGGATTTATAAAATTTTGGAACTAAACGGGTACGATCCCCAATATGCGGCT 1259 TCGAGTAATCGGATTTACAAAATCCTCGAACTTAACGGATACGACCCCCAATACGCCGCT TCTTCTAACCGTATCTACAAGATCCTTGAGTTGAATGGCTACGACCCACAGTACGCCGCC ** ** ** ** ** ** ** * ** * ** ** ***** ** ** ** ** **
WTREP dREP sREP
TCCGTCTTTCTGGGATGGGCCACGAAAAAGTTCGGCAAGAGGAACACCATCTGGCTGTTT 1319 AGCGTGTTTCTGGGGTGGGCGACTAAGAAATTCGGAAAGCGTAATACGATTTGGTTGTTC TCAGTGTTTCTGGGCTGGGCAACCAAGAAATTTGGGAAACGCAATACGATTTGGCTGTTC ** ******** ***** ** ** ** ** ** ** * ** ** ** *** ****
WTREP dREP sREP
GGGCCTGCAACTACCGGGAAGACCAACATCGCGGAGGCCATAGCCCACACTGTGCCCTTC 1379 GGACCCGCTACGACCGGCAAAACGAATATCGCCGAAGCGATCGCGCATACCGTGCCATTC GGACCCGCCACCACTGGTAAGACTAATATTGCCGAGGCGATCGCACATACCGTTCCGTTT ** ** ** ** ** ** ** ** ** ** ** ** ** ** ** ** ** ** ** **
WTREP dREP sREP
TACGGGTGCGTAAACTGGACCAATGAGAACTTTCCCTTCAACGACTGTGTCGACAAGATG 1439 TACGGGTGCGTGAATTGGACGAACGAGAACTTTCCGTTTAACGATTGCGTCGACAAGATG TACGGATGCGTGAATTGGACTAACGAAAATTTCCCCTTTAATGATTGCGTGGACAAGATG ***** ***** ** ***** ** ** ** ** ** ** ** ** ** ** *********
WTREP dREP sREP
GTGATCTGGTGGGAGGAGGGGAAGATGACCGCCAAGGTCGTGGAGTCGGCCAAAGCCATT 1499 GTGATTTGGTGGGAGGAGGGAAAGATGACCGCTAAGGTGGTCGAGTCCGCGAAAGCGATT GTTATTTGGTGGGAGGAAGGAAAGATGACTGCGAAAGTGGTGGAATCCGCTAAGGCTATC ** ** *********** ** ******** ** ** ** ** ** ** ** ** ** **
WTREP dREP sREP
CTCGGAGGAAGCAAGGTGCGCGTGGACCAGAAATGCAAGTCCTCGGCCCAGATAGACCCG 1559 CTGGGGGGGTCTAAGGTGAGAGTCGACCAGAAGTGTAAGTCTTCGGCTCAGATCGATCCG TTGGGGGGGTCGAAAGTTCGGGTCGACCAGAAGTGCAAATCGTCCGCGCAGATTGACCCC * ** ** ** ** * ** ******** ** ** ** ** ** ***** ** **
WTREP dREP sREP
ACTCCCGTGATCGTCACCTCCAACACCAACATGTGCGCCGTGATTGACGGGAACTCAACG 1619 ACCCCCGTGATCGTGACCTCTAACACTAACATGTGCGCCGTGATCGACGGGAATTCGACT ACCCCCGTGATTGTGACGTCAAATACTAATATGTGTGCGGTCATCGATGGCAATAGCACC ** ******** ** ** ** ** ** ** ***** ** ** ** ** ** ** **
WTREP dREP sREP
ACCTTCGAACACCAGCAGCCGTTGCAAGACCGGATGTTCAAATTTGAACTCACCCGCCGT 1679 ACGTTCGAACACCAGCAGCCATTGCAGGACCGTATGTTCAAATTTGAACTGACTAGGAGA ACTTTCGAACATCAGCAACCCCTCCAGGATCGTATGTTTAAGTTCGAGTTGACTCGGCGG ** ******** ***** ** * ** ** ** ***** ** ** ** * ** * *
WTREP
CTGGATCATGACTTTGGGAAGGTCACCAAGCAGGAAGTCAAAGACTTTTTCCGGTGGGCA 1739
(Continued)
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dREP sREP
WTREP dREP sREP
WTREP dREP sREP
CTCGACCACGACTTCGGAAAGGTGACTAAGCAGGAGGTGAAAGACTTTTTTCGGTGGGCG CTGGACCACGATTTCGGCAAAGTGACGAAACAGGAGGTGAAGGACTTCTTTAGATGGGCC ** ** ** ** ** ** ** ** ** ** ***** ** ** ***** ** * ***** GGT AAGGATCACGTGGTTGAGGTGGAGCATGAATTCTACGTCAAAAAGGGTGGAGCCAAGAAA 1799 AAAGACCATGTGGTCGAGGTCGAGCACGAGTTTTACGTGAAAAAGGGCGGAGCGAAAAAG AAGGACCACGTGGTGGAGGTCGAGCACGAGTTTTATGTGAAGAAGGGGGGGGCCAAAAAG ** ** ** ***** ***** ***** ** ** ** ** ** ***** ** ** ** ** SP1 AP1 AGACCCGCCCCCAGTGACGCAGATATAAGTGAGCCCAAACGGGTGCGCGAGTCAGTTGCG 1859 AGACCCGCCCCTAGCGACGCCGACATTAGCGAACCGAAACGCGTACGCGAATCCGTTGCG CGCCCTGCACCTTCCGACGCCGACATTTCCGAGCCAAAGAGAGTGCGTGAGAGTGTGGCC * ** ** ** ***** ** ** ** ** ** * ** ** ** ** **
WTREP dREP sREP
CAGCCATCGACGTCAGACGCGGAAGCTTCGATCAACTACGCAGACAGGTACCAAAACAAA 1919 CAACCGTCAACCTCCGACGCCGAAGCGTCAATCAATTACGCCGATAGGTACCAGAATAAG CAACCCTCCACCAGTGATGCCGAGGCCTCCATTAATTATGCCGACCGCTATCAGAATAAG ** ** ** ** ** ** ** ** ** ** ** ** ** ** * ** ** ** **
WTREP dREP sREP
TGTTCTCGTCACGTGGGCATGAATCTGATGCTGTTTCCCTGCAGACAATGCGAGAGAATG 1979 TGCTCTAGACACGTGGGGATGAATCTGATGCTGTTTCCCTGTAGACAGTGCGAGCGTATG TGCTCAAGGCATGTCGGGATGAACCTGATGCTGTTCCCATGCCGCCAGTGCGAGCGCATG ** ** * ** ** ** ***** *********** ** ** * ** ****** * ***
WTREP dREP sREP
AATCAGAATTCAAATATCTGCTTCACTCACGGACAGAAAGACTGTTTAGAGTGCTTTCCC 2039 AACCAGAACTCGAACATTTGCTTTACCCACGGACAGAAAGACTGTCTCGAATGCTTTCCC AACCAGAACAGCAACATTTGTTTTACCCACGGGCAGAAGGATTGCCTGGAATGCTTCCCG ** ***** ** ** ** ** ** ***** ***** ** ** * ** ***** **
WTREP dREP sREP
GTGTCAGAATCTCAACCCGTTTCTGTCGTCAAAAAGGCGTATCAGAAACTGTGCTACATT 2099 GTGTCCGAATCGCAACCCGTTAGCGTGGTGAAAAAAGCGTACCAGAAACTGTGTTACATA GTCAGCGAGTCACAGCCGGTGTCCGTGGTGAAGAAAGCCTACCAAAAGCTGTGTTACATC ** ** ** ** ** ** ** ** ** ** ** ** ** ** ***** *****
WTREP dREP sREP
CATCATATCATGGGAAAGGTGCCAGACGCTTGCACTGCCTGCGATCTGGTCAATGTGGAT 2159 CACCATATTATGGGCAAAGTGCCCGACGCATGCACCGCATGCGATCTGGTGAACGTCGAC CACCACATTATGGGGAAAGTCCCCGATGCCTGTACCGCATGCGACCTGGTGAACGTTGAC ** ** ** ***** ** ** ** ** ** ** ** ** ***** ***** ** ** **
WTREP dREP sREP
TTGGATGACTGCATCTTTGAACAATAA CTCGACGATTGCATTTTTGAACAGTAA CTCGACGACTGCATTTTCGAGCAGTAA * ** ** ***** ** ** ** ***
2186
Fig. S1. Alignments of WT, deoptimized (d), or scrambled (s) AAV Rep. Alignments of the 1,866-bp recoded Rep genomic sequences were completed and displayed by using Clustal 2.1 multiple sequence alignment tool, numbered relative to the full-length 4,679-bp AAV2 genome (GenBank accession no. AF043303.1); nucleotides identical to all three sequences are delineated by asterisks. The black dot delineates the initiator MET starting at base pair 321, the p19 (yellow) and p40 (green) promoter TATA sequences are highlighted, and the 5′-Rep 68/40 alternative splice site is delineated in bold/underlined. The 135bp Rep inhibitory sequences encompassing nucleotides 1,782 to 1,916 are delineated in red (refer to text for details), and previously characterized p40 promoter transcription binding sites for GGT, SP1, and AP1 (boxed) are shown; the transcription start site is delineated (arrow).
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Fig. S2. Functional characterization of single-backbone Ad/AAV/dRep hybrid virus. (A) Schema of Ad/AAV/dRep with predicted monomeric (M) and dimeric (D) forms resulting from productive Rep 78 cleavage of endogenous AAV integrating elements p5IEE and AAV TR (Fig. 2 shows excision model). Sizes are as follows: ψ, Ad packaging sequence (0.4 kb); pCMV/EGFP cassette (1.6 kb); pTK/NeoR, (1.1 kb); Ad, Ad base pairs 3,330 to 3,940 (0.6 kb); p5IEE, 135 bp; AAV TR (145 bp each plus G–C tail); and dRep, approximately 1.9 kb. (B) Ad/AAV/dRep was serially passaged in 293 cells, and at indicated passages (P), Hirt DNA was isolated for genomic analysis using the approximately 0.2-kb Ad left end inverted TR (ITR base pairs 1–194) as probe (horizontal bar). *Parent Ad/AAV/dRep virus.
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Start (BP) 321
ATGCCCGGATTCTACGAAATCGTCATCAAAGTGCCCTCTGACTTGGATGAACACCTGCCGGGGATCAGCGATTCTTTCGTCAATTGGGTCGCGGAGAAAGAGTGGGAACTTCCCCCCGACTCGGACATGGAC
End (BP) 452
453
CTGAACTTAATCGAGCAAGCCCCGCTGACGGTGGCGGAGAAACTGCAGCGGGACTTTCTGACCGAGTGGAGGCGCGTATCGAAAGCGCCCGAAGCTTTGTTTTTCGTCCAGTTCGAGAAGGGGGAGTCGTAC
584
585
TTTCATATGCATGTGTTGGTGGAGACTACGGGAGTGAAGAGTATGGTGCTAGGGAGGTTTCTGTCGCAAATAAGAGAGAAGCTGATCCAGCGGATATACCGTGGCATTGAGCCCACCCTTCCCAATTGGTTT
716
717
GCCGTGACCAAAACTCGTAACGGAGCAGGGGGGGGAAATAAAGTCGTCGACGAGTGCTATATTCCGAACTACCTCTTGCCCAAGACGCAGCCCGAATTGCAGTGGGCCTGGACCAACATGGAGCAATACCTG
848
849
TCAGCGTGCCTCAACTTGACCGAAAGAAAGAGACTCGTGGCCCAGCACCTGACCCATGTCTCACAGACCCAGGAACAGAATAAGGAAAACCAAAACCCAAATAGCGACGCCCCCGTGATACGGAGCAAGACC
981
AGCGCTCGCTACATGGAGTTAGTGGGATGGTTGGTGGATAAAGGAATCACGTCTGAGAAACAATGGATTCAGGAGGACCAGGCGTCCTACATTAGTTTTAACGCCGCGTCAAATAGCAGATCTCAGATTAAA
1113
GCCGCGCTCGATAACGCCGGCAAAATCATGTCGCTGACCAAGACAGCTCCCGACTACCTGGTGGGACAGCAGCCGGTGGAGGACATCTCTTCTAACCGTATCTACAAGATCCTTGAGTTGAATGGCTACGAC
1244
1245
CCCCAATATGCGGCTTCCGTCTTTCTGGGATGGGCCACGAAAAAGTTCGGCAAGAGGAACACCATCTGGCTGTTTGGGCCTGCAACTACCGGGAAGACCAACATCGCGGAGGCCATAGCCCACACTGTGCCC
1376
1377
TTCTACGGGTGCGTAAACTGGACCAATGAGAACTTTCCCTTCAACGACTGTGTCGACAAGATGGTGATCTGGTGGGAGGAGGGGAAGATGACCGCCAAGGTCGTGGAGTCGGCCAAAGCCATTCTCGGAGGAAGC
1511
1512
AAGGTGCGCGTGGACCAGAAATGCAAGTCCTCGGCCCAGATAGACCCGACTCCCGTGATCGTCACCTCCAACACCAACATGTGCGCCGTGATTGACGGGAACTCAACGACCTTCGAACACCAGCAGCCGTTGCAA 1646
1647
GACCGGATGTTCAAATTTGAACTCACCCGCCGTCTGGATCATGACTTTGGGAAGGTCACCAAGCAGGAAGTCAAAGACTTTTTCCGGTGGGCAAAGGATCACGTGGTTGAGGTGGAGCATGAATTCTACGTCAAA
1781
1782
AAGGGTGGAGCCAAGAAAAGACCCGCCCCCAGTGACGCAGATATAAGTGAGCCCAAACGGGTGCGCGAGTCAGTTGCGCAGCCATCGACGTCAGACGCGGAAGCTTCGATCAACTACGCAGACAGGTACCAAAAC
1916
1917
AAATGTTCTCGTCACGTGGGCATGAATCTGATGCTGTTTCCCTGCAGACAATGCGAGAGAATGAATCAGAATTCAAATATCTGCTTCACTCACGGACAGAAAGACTGTTTAGAGTGCTTTCCCGTGTCAGAATCT 2051
2052
CAACCCGTTTCTGTCGTCAAAAAGGCGTATCAGAAACTGTGCTACATTCATCATATCATGGGAAAGGTGCCAGACGCTTGCACTGCCTGCGATCTGGTCAATGTGGATTTGGATGACTGCATCTTTGAACAATAA
Rep78-I: SSSSSSSWWWWWWW †
980 1112
2186
Start (BP) 321
ATGCCCGGATTCTACGAAATCGTCATCAAAGTGCCCTCTGACTTGGATGAACACCTGCCGGGGATCAGCGATTCTTTCGTCAATTGGGTCGCGGAGAAAGAGTGGGAACTTCCCCCCGACTCGGACATGGAC
End (BP) 452
453
CTGAACTTAATCGAGCAAGCCCCGCTGACGGTGGCGGAGAAACTGCAGCGGGACTTTCTGACCGAGTGGAGGCGCGTATCGAAAGCGCCCGAAGCTTTGTTTTTCGTCCAGTTCGAGAAGGGGGAGTCGTAC
584
585
TTTCATATGCATGTGTTGGTGGAGACTACGGGAGTGAAGAGTATGGTGCTAGGGAGGTTTCTGTCGCAAATAAGAGAGAAGCTGATCCAGCGGATATACCGTGGCATTGAGCCCACCCTTCCCAATTGGTTT
716
717
GCGGTCACAAAGACCAGAAATGGCGCCGGAGGCGGGAACAAGGTGGTGGATGAGTGCTACATCCCCAATTACTTGCTCCCCAAAACCCAGCCTGAGCTCCAGTGGGCGTGGACTAATATGGAACAGTATTTA
848
849
AGCGCCTGTTTGAATCTCACGGAGCGTAAACGGTTGGTGGCGCAGCATCTGACGCAGTGTCGCAGACGCAGGAGCAGAACAAAGAGAATCAGAATCCCAATTCTGATGCGCCGGTGATCAGATCAAAAACT
980
981
TCAGCCAGGTACATGGAGCTGGTCGGGTGGCTCGTGGACAAGGGGATTACCTCGGAGAAGCAGTGGATCCAGGAGGACCAGGCCTCATACATCTCCTTCAATGCGGCCTCCAACTCGCGGTCCCAAATCAAG
1112
1113
GCTGCCTTGGACAATGCGGGAAAGATTATGAGCCTGACTAAAACCGCCCCCGACTACCTGGTGGGCCAGCAGCCCGTGGAGGACATTTCCAGCAATCGGATTTATAAAATTTTGGAACTAAACGGGTACGAT
1244
1245
CCCCAATATGCGGCTTCCGTCTTTCTGGGATGGGCCACGAAAAAGTTCGGCAAGAGGAACACCATCTGGCTGTTTGGGCCTGCAACTACCGGGAAGACCAACATCGCGGAGGCCATAGCCCACACTGTGCCC
1376
1377
TTCTACGGGTGCGTAAACTGGACCAATGAGAACTTTCCCTTCAACGACTGTGTCGACAAGATGGTGATCTGGTGGGAGGAGGGGAAGATGACCGCCAAGGTCGTGGAGTCGGCCAAAGCCATTCTCGGAGGAAGC
1511
1512
AAGGTGCGCGTGGACCAGAAATGCAAGTCCTCGGCCCAGATAGACCCGACTCCCGTGATCGTCACCTCCAACACCAACATGTGCGCCGTGATTGACGGGAACTCAACGACCTTCGAACACCAGCAGCCGTTGCAA
1646
1647
GATCGTATGTTTAAGTTCGAGTTGACTCGGCGGCTGGACCACGATTTCGGCAAAGTGACGAAACAGGAGGTGAAGGACTTCTTTAGATGGGCCAAGGACCACGTGGTGGAGGTCGAGCACGAGTTTTATGTGAAG
1781
1782
AAGGGGGGGGCCAAAAAGCGCCCTGCACCTTCCGACGCCGACATTTCCGAGCCAAAGAGAGTGCGTGAGAGTGTGGCCCAACCCTCCACCAGTGATGCCGAGGCCTCCATTAATTATGCCGACCGCTATCAGAAT
1916
1917
AAGTGCTCAAGGCATGTCGGGATGAACCTGATGCTGTTCCCATGCCGCCAGTGCGAGCGCATGAACCAGAACAGCAACATTTGTTTTACCCACGGGCAGAAGGATTGCCTGGAATGCTTCCCGGTCAGCGAGTCA
2051
2052
CAGCCGGTGTCCGTGGTGAAGAAAGCCTACCAAAAGCTGTGTTACATCCACCACATTATGGGGAAAGTCCCCGATGCCTGTACCGCATGCGACCTGGTGAACGTTGACCTCGACGACTGCATTTTCGAGCAGTAA
2186
Rep78-II: SSSWWWWWWWSSSS †
Start
End
321
Rep78- III: WWSSSWWWSSSWW † ATGCCGGGGTTTTACGAGATTGTGATTAAGGTCCCCAGCGACCTTGACGAGCATCTGCCCGGCATTTCTGACAGCTTTGTGAACTGGGTGGCCGAGAAGGAATGGGAGTTGCCGCCAGATTCTGACATGGAT
452
453
CTGAATCTGATTGAGCAGGCACCCCTGACCGTGGCCGAGAAGCTGCAGCGCGACTTTCTGACGGAATGGCGCCGTGTGAGTAAGGCCCCGGAGGCCCTTTTCTTTGTGCAATTTGAGAAGGGAGAGAGCTAC
584
585
TTTCATATGCATGTGTTGGTGGAGACTACGGGAGTGAAGAGTATGGTGCTAGGGAGGTTTCTGTCGCAAATAAGAGAGAAGCTGATCCAGCGGATATACCGTGGCATTGAGCCCACCCTTCCCAATTGGTTT
716
717
GCCGTGACCAAAACTCGTAACGGAGCAGGGGGGGGAAATAAAGTCGTCGACGAGTGCTATATTCCGAACTACCTCTTGCCCAAGACGCAGCCCGAATTGCAGTGGGCCTGGACCAACATGGAGCAATACCTG
848
849
TCAGCGTGCCTCAACTTGACCGAAAGAAAGAGACTCGTGGCCCAGCACCTGACCCATGTCTCACAGACCCAGGAACAGAATAAGGAAAACCAAAACCCAAATAGCGACGCCCCCGTGATACGGAGCAAGACC
980
981
TCAGCCAGGTACATGGAGCTGGTCGGGTGGCTCGTGGACAAGGGGATTACCTCGGAGAAGCAGTGGATCCAGGAGGACCAGGCCTCATACATCTCCTTCAATGCGGCCTCCAACTCGCGGTCCCAAATCAAG
1112
1113
GCTGCCTTGGACAATGCGGGAAAGATTATGAGCCTGACTAAAACCGCCCCCGACTACCTGGTGGGCCAGCAGCCCGTGGAGGACATTTCCAGCAATCGGATTTATAAAATTTTGGAACTAAACGGGTACGAT
1244
1245
CCCCAATATGCGGCTTCCGTCTTTCTGGGATGGGCCACGAAAAAGTTCGGCAAGAGGAACACCATCTGGCTGTTTGGGCCTGCAACTACCGGGAAGACCAACATCGCGGAGGCCATAGCCCACACTGTGCCC
1376
1377
TTTTACGGATGCGTGAATTGGACTAACGAAAATTTCCCCTTTAATGATTGCGTGGACAAGATGGTTATTTGGTGGGAGGAAGGAAAGATGACTGCGAAAGTGGTGGAATCCGCTAAGGCTATCTTGGGGGGGTCG
1511
1512
AAAGTTCGGGTCGACCAGAAGTGCAAATCGTCCGCGCAGATTGACCCCACCCCCGTGATTGTGACGTCAAATACTAATATGTGTGCGGTCATCGATGGCAATAGCACCACTTTCGAACATCAGCAACCCCTCCAG
1646
1647
GATCGTATGTTTAAGTTCGAGTTGACTCGGCGGCTGGACCACGATTTCGGCAAAGTGACGAAACAGGAGGTGAAGGACTTCTTTAGATGGGCCAAGGACCACGTGGTGGAGGTCGAGCACGAGTTTTATGTGAAG
1781
1782
AAGGGGGGGGCCAAAAAGCGCCCTGCACCTTCCGACGCCGACATTTCCGAGCCAAAGAGAGTGCGTGAGAGTGTGGCCCAACCCTCCACCAGTGATGCCGAGGCCTCCATTAATTATGCCGACCGCTATCAGAAT
1916
1917
AAATGTTCTCGTCACGTGGGCATGAATCTGATGCTGTTTCCCTGCAGACAATGCGAGAGAATGAATCAGAATTCAAATATCTGCTTCACTCACGGACAGAAAGACTGTTTAGAGTGCTTTCCCGTGTCAGAATCT
2051
2052
CAACCCGTTTCTGTCGTCAAAAAGGCGTATCAGAAACTGTGCTACATTCATCATATCATGGGAAAGGTGCCAGACGCTTGCACTGCCTGCGATCTGGTCAATGTGGATTTGGATGACTGCATCTTTGAACAATAA
2186
(BP)
(BP)
Start (BP) 321
ATGCCGGGGTTTTACGAGATTGTGATTAAGGTCCCCAGCGACCTTGACGAGCATCTGCCCGGCATTTCTGACAGCTTTGTGAACTGGGTGGCCGAGAAGGAATGGGAGTTGCCGCCAGATTCTGACATGGAT
End (BP) 452
453
CTGAACTTAATCGAGCAAGCCCCGCTGACGGTGGCGGAGAAACTGCAGCGGGACTTTCTGACCGAGTGGAGGCGCGTATCGAAAGCGCCCGAAGCTTTGTTTTTCGTCCAGTTCGAGAAGGGGGAGTCGTAC
584
585
TTCCACATGCACGTGCTCGTGGAAACCACCGGGGTGAAATCCATGGTTTTGGGACGTTTCCTGAGTCAGATTCGCGAAAAACTGATTCAGAGAATTTACCGCGGGATCGAGCCGACTTTGCCAAACTGGTTC
716
717
GCGGTCACAAAGACCAGAAATGGCGCCGGAGGCGGGAACAAGGTGGTGGATGAGTGCTACATCCCCAATTACTTGCTCCCCAAAACCCAGCCTGAGCTCCAGTGGGCGTGGACTAATATGGAACAGTATTTA
848
849
TCAGCGTGCCTCAACTTGACCGAAAGAAAGAGACTCGTGGCCCAGCACCTGACCCATGTCTCACAGACCCAGGAACAGAATAAGGAAAACCAAAACCCAAATAGCGACGCCCCCGTGATACGGAGCAAGACC
980
981
AGCGCTCGCTACATGGAGTTAGTGGGATGGTTGGTGGATAAAGGAATCACGTCTGAGAAACAATGGATTCAGGAGGACCAGGCGTCCTACATTAGTTTTAACGCCGCGTCAAATAGCAGATCTCAGATTAAA
1112
1113
GCTGCCTTGGACAATGCGGGAAAGATTATGAGCCTGACTAAAACCGCCCCCGACTACCTGGTGGGCCAGCAGCCCGTGGAGGACATTTCCAGCAATCGGATTTATAAAATTTTGGAACTAAACGGGTACGAT
1244
1245
CCACAGTACGCCGCCTCAGTGTTTCTGGGCTGGGCAACCAAGAAATTTGGGAAACGCAATACGATTTGGCTGTTCGGACCCGCCACCACTGGTAAGACTAATATTGCCGAGGCGATCGCACATACCGTTCCG
1376
1377
TTTTACGGATGCGTGAATTGGACTAACGAAAATTTCCCCTTTAATGATTGCGTGGACAAGATGGTTATTTGGTGGGAGGAAGGAAAGATGACTGCGAAAGTGGTGGAATCCGCTAAGGCTATCTTGGGGGGGTCG
1511
1512
AAGGTGCGCGTGGACCAGAAATGCAAGTCCTCGGCCCAGATAGACCCGACTCCCGTGATCGTCACCTCCAACACCAACATGTGCGCCGTGATTGACGGGAACTCAACGACCTTCGAACACCAGCAGCCGTTGCAA
1646
1647
GACCGGATGTTCAAATTTGAACTCACCCGCCGTCTGGATCATGACTTTGGGAAGGTCACCAAGCAGGAAGTCAAAGACTTTTTCCGGTGGGCAAAGGATCACGTGGTTGAGGTGGAGCATGAATTCTACGTCAA
1781
1782
AAGGGGGGGGCCAAAAAGCGCCCTGCACCTTCCGACGCCGACATTTCCGAGCCAAAGAGAGTGCGTGAGAGTGTGGCCCAACCCTCCACCAGTGATGCCGAGGCCTCCATTAATTATGCCGACCGCTATCAGAAT
1916
1917
AAGTGCTCAAGGCATGTCGGGATGAACCTGATGCTGTTCCCATGCCGCCAGTGCGAGCGCATGAACCAGAACAGCAACATTTGTTTTACCCACGGGCAGAAGGATTGCCTGGAATGCTTCCCGGTCAGCGAGTCA
2051
2052
CAACCCGTTTCTGTCGTCAAAAAGGCGTATCAGAAACTGTGCTACATTCATCATATCATGGGAAAGGTGCCAGACGCTTGCACTGCCTGCGATCTGGTCAATGTGGATTTGGATGACTGCATCTTTGAACAATAA
2186
Rep78- IV: WSWWSSWSSWWSSW †
Fig. S3. Nucleotide sequences of Rep chimers used for predictive combinatorial group testing. WT (W) and scrambled (S; italicized) sequences of the four mutant Rep genes are displayed. The segment encompassing the critical inhibitory domain in Rep 78-I (base pairs 1,782–1,916) is highlighted in red.
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Table S1.
Oligonucleotides used for qPCR
Primer AdenoqPCR-1 AdenoqPCR-2 B-ActinF B-ActinR
Sequence
Nucleotide position
Accession no.
Gene ID
5′-CTGTGATGCTGGATGTGACC-3′ 5′-TGCTTCCATCAAACGAGTTG-3′ 5′-TCACCCACACTGTGCCCATCTACGA-3′ 5′-CAGCGGAACCGCTCATTGCCAATGG-3′
3,412–3,431 3,636–3,617 560–584 854–830
GB AY339865.1
Human Ad C serotype 5 genome
NM_001101.3
β-Actin mRNA, human
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