LOCUS pHACK-GAL4_p65AD 10248 bp DNA circular SYN 08-MAR-2022 DEFINITION . ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM recombinant plasmid REFERENCE 1 (bases 1 to 10248) AUTHORS Junjie Luo TITLE Direct Submission JOURNAL Exported Apr 26, 2025 from SnapGene 8.0.3 https://www.snapgene.com FEATURES Location/Qualifiers source 1..10248 /mol_type="other DNA" /organism="recombinant plasmid" protein_bind 41..74 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" /note="color: #31849b; direction: RIGHT" protein_bind 88..157 /label=attB /bound_moiety="phage phi-C31 integrase" /note="attB site for the phi-C31 integrase (Groth et al., 2000)" /note="color: #31849b" protein_bind complement(88..157) /label=attB /bound_moiety="phage phi-C31 integrase" /note="attB site for the phi-C31 integrase (Groth et al., 2000)" /note="color: #31849b; direction: LEFT" primer_bind complement(88..110) /label=AD2_attB_typing_f2 /note="color: black; sequence: GCGCGTACTCCACCTCACCCATC; added: 2020-02-17" primer_bind complement(137..159) /label=GFP_attB_f /note="color: black; sequence: aaactcatcaatgtatcttaCTCGAAGCCGCGGTGCGGGTG" primer_bind complement(137..157) /label=AD2_attB_typing_f /note="color: black; sequence: CTCGAAGCCGCGGTGCGGGTG; added: 2020-02-17" misc_feature join(175..205,206..313) /label=P element 5' end (Potter) /note="first 139 bp of P element 5' end (586 bp)" /note="This feature has 2 segments: 1: 175 .. 205 / #ffcc99 / inverted repreat 2: 206 .. 313 / #ffcc99" protein_bind 326..359 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" /note="color: #31849b; direction: RIGHT" misc_feature 399..1555 /label=GAL4_HA1 /note="color: #a6acb3; direction: RIGHT" CDS 1568..1621 /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /note="color: #cc99b2 Cleavage site after base 1618" /translation="EGRGSLLTCGDVEENPGP" primer_bind 1620..1654 /label=p65ADZp_f /note="color: black; sequence: CCCCtaggATGGATAAAGCGGAATTAATTCCCGAG; added: 2022-03-07" misc_feature 1623..2993 /label=Insertion /note="color: #ff0000" CDS 1628..2686 /codon_start=1 /label=p65ADZp /label=ORF frame 1 /note="color: #993366" /translation="MDKAELIPEPPKKKRKVELGSTPMEFQYLPDTDDRHRIEEKRKRT YETFKSIMKKSPFSGPTDPRPPPRRIAVPSRSSASVPKPAPQPYPFTSSLSTINYDEFP TMVFPSGQISQASALAPAPPQVLPQAPAPAPAPAMVSALAQAPAPVPVLAPGPPQAVAP PAPKPTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNSEFQQLLNQGI PVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGDEDFSSIADMD FSALLSQISSASGGGGGGGGGGTSLEIEAAFLERENTALETRVAELRQRVQRLRNRVSQ YRTRYGPLGGGK" misc_feature 1637..1684 /label=SV40_NLS /note="color: #a6acb3" CDS 1658..1678 /codon_start=1 /product="nuclear localization signal of SV40 large T antigen" /label=SV40 NLS /note="color: #993366" /translation="PKKKRKV" primer_bind complement(2519..2536) /label=Zp_primer_r /note="color: black; sequence: ACGGTATCGATAGACGTCACCTCCACCTCCACCTCC; added: 2020-12-27" polyA_signal 2702..2947 /label=hsp70 polyA /note="color: #a6acb3; direction: RIGHT" primer_bind complement(2923..2997) /label=p65ADZp_LoxP_r /note="color: black; sequence: GGCATATGataacttcgtataatgtatgctatacgaagttatgcggccgcGATCTAAAC GAGTTTTTAAGCAAAC; added: 2022-03-07" protein_bind 2956..2989 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." /note="color: #31849b; direction: RIGHT" misc_feature join(2994,2995..3044,3045..3050,3051..4210,4211..4227) /label=[AD2_B^{Bxb1}] /note="This forward directional feature has 5 segments: 1: 2994 .. 2994 / #ffffff 2: 2995 .. 3044 / #31849b / attB[Bxb1] 3: 3045 .. 3050 / #ffffff 4: 3051 .. 4210 / #00ffff / 2xr4-CFP 5: 4211 .. 4227 / #ffffff" misc_feature complement(2995..3044) /label=attB(Bxb1) /note="http://flybase.org/reports/FBto0000359 The attB(Bxb1) site (5'-TCGGCCGGCTTGTCGACGACGgcggtctcCGTCGTCAGGATCATCCGGGC-3') is one of the two target sites of the Bxb1:int integrase. Bxb1:int mediates recombination between two non-identical target sites, attP(Bxb1) and attB(Bxb1), each of which contains a short integration core flanked by imperfect inverted repeats. Exchange between the attP(Bxb1)/attB(Bxb1) pair results in the formation of hybrid sites that are no longer a substrate of the Bxb1:int integrase, and thus the exchange reaction driven by Bxb1:int is undirectional (reviewed in PMID:21195181, FBrf0231034). The recombination event catalyzed by Bxb1:int results in genetic modification, the nature of which is influenced by the relative orientation (direct or inverted), and location of the two target sites. The types of possible modification include deletion of DNA, generation of chromosomal rearrangements, integration of DNA into the genome, and replacement of genetic material with that from a donor plasmid using recombination-mediated cassette exchange (RMCE) (reviewed in FBrf0231034)." /note="color: #31849b; direction: LEFT" polyA_signal 3051..3172 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" /note="color: #a6acb3" primer_bind complement(3051..3070) /label=GFP_attB_f /note="color: black; sequence: aaactcatcaatgtatcttaCTCGAAGCCGCGGTGCGGGTG" primer_bind complement(3067..3096) /label=SV40A_seq_f /note="color: black; sequence: CACTGCATTCTAGTTGTGGTTTGTCCAAAC; added: 2021-10-19" CDS complement(join(3179..3892,3893..3895,3896..3898)) /codon_start=1 /product="cyan variant of GFP" /label=CFP /note="mammalian codon-optimized" /note="This feature has 3 segments: 1: 3179 .. 3892 / #00ffff 2: 3893 .. 3895 / #00ffff / 1a 3: 3896 .. 3898 / #00ffff" /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(3182..3211) /label=CFP_230_seq_fL /note="color: black; sequence: ATCACTCTCGGCATGGACGAGCTGTACAAG; added: 2021-12-14" primer_bind complement(3182..3204) /label=CFP_230_seq_f /note="color: black; sequence: TCGGCATGGACGAGCTGTACAAG; added: 2021-09-18" primer_bind 3761..3783 /label=CFP_45_seq_r /note="color: black; sequence: CTTCAGGGTCAGCTTGCCGTAGG; added: 2021-09-18" promoter complement(join(3909..3994,4022..4027,4028..4033)) /label=hsp70 mini /note="Drosophila melanogaster hsp70Bb promoter" /note="This reverse directional feature has 3 segments: 1: 3909 .. 3994 / #ffffff / align to 3xP3 2: 4022 .. 4027 / #cc99ff / TATA box 3: 4028 .. 4033 / #ffffff" promoter complement(join(4053..4068,4073..4088,4093..4108, 4113..4128)) /label=r4 promoter /note="This reverse directional feature has 4 segments: 1: 4053 .. 4068 / #ffffff 2: 4073 .. 4088 / #ffffff 3: 4093 .. 4108 / #ffffff 4: 4113 .. 4128 / #ffffff" promoter complement(join(4135..4150,4155..4170,4175..4190, 4195..4210)) /label=r4 promoter /note="This reverse directional feature has 4 segments: 1: 4135 .. 4150 / #ffffff 2: 4155 .. 4170 / #ffffff 3: 4175 .. 4190 / #ffffff 4: 4195 .. 4210 / #ffffff" protein_bind 4233..4266 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." /note="color: #31849b; direction: RIGHT" misc_feature 4277..5645 /label=GAL4_HA2 /note="color: #a6acb3; direction: RIGHT" CDS 5301..5645 /codon_start=1 /gene="Saccharomyces cerevisiae GAL4 (truncated)" /product="activation domain of the GAL4 transcriptional activator" /label=GAL4 activation domain /note="color: #993366" /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD EDTPPNPKKE" protein_bind 5655..5688 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" /note="color: #31849b; direction: RIGHT" promoter 5704..6500 /label=dU6-1 promoter /note="RNA polymerase III promoter for Drosophila U6-1 snRNA (Port et al., 2014)" /note="color: #ffffff; direction: RIGHT" primer_bind 5965..5975 /label=Zp_primer_r /note="color: black; sequence: ACGGTATCGATAGACGTCACCTCCACCTCCACCTCC; added: 2020-12-27" misc_RNA 6520..6595 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" /note="color: #00ccff; direction: RIGHT" promoter 6596..7016 /label=dU6-3 promoter /note="RNA polymerase III promoter for Drosophila U6-3 snRNA (Port et al., 2014)" /note="color: #ffffff; direction: RIGHT" promoter 6602..7016 /label=U6:3 promoter /note="color: #ffffff; direction: RIGHT" misc_RNA 7037..7112 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" /note="color: #00ccff; direction: RIGHT" misc_feature join(7372..7503,7504..7534) /label=P element 3' end (Potter) /note="last 163 bp of P element 3' end (233 bp) Editing Transgenic DNA Components by Inducible Gene Replacement in Drosophila melanogaster Editing Transgenic DNA Components by InducibleGene Replacement in Drosophila melanogaster Chun-Chieh Lin and Christopher J. Potter1 The Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 Editing Transgenic DNA Components by Inducible Gene Replacement in Drosophila melanogaster To generate a versatile construct with multiple synthetic com-ponents, we de novo designed the donor vector for DNA syn-thesis (DNA2.0, Newark, CA). The construct contains 59 and 39P-element sequences, attB sequence, T2A, origin of replication,and ampicillin resistance sequence. " /note="This feature has 2 segments: 1: 7372 .. 7503 / #ffcc99 2: 7504 .. 7534 / #ffcc99 / inverted repeat" protein_bind complement(7545..7578) /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" /note="color: #31849b; direction: LEFT" primer_bind complement(8048..8065) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" /note="color: black; sequence: TGTAAAACGACGGCCAGT" primer_bind complement(8048..8064) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" /note="color: #a020f0; direction: LEFT" primer_bind complement(8163..8180) /label=L4440 /note="L4440 vector, forward primer" /note="color: black; sequence: AGCGAGTCAGTGAGCGAG" rep_origin complement(8334..8922) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" /note="color: #ffff00" primer_bind complement(8414..8433) /label=pBR322ori-F /note="pBR322 origin, forward primer" /note="color: black; sequence: GGGAAACGCCTGGTATCTTT" CDS complement(join(9044..9835,9836..9904)) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /note="This feature has 2 segments: 1: 9044 .. 9835 / #ccffcc 2: 9836 .. 9904 / #ccffcc / signal sequence Cleavage site after base 9835" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 9667..9686 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" /note="color: black; sequence: ATAATACCGCGCCACATAGC" promoter complement(9905..9996) /gene="bla" /label=AmpR promoter /note="color: #ffffff; direction: LEFT" primer_bind 10038..10057 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" /note="color: black; sequence: ATCGCGAGCCCATTTATACC" primer_bind 10063..10082 /label=pENTR-R /note="pENTR vectors, reverse primer" /note="color: black; sequence: ATGGCTCATAACACCCCTTG" ORIGIN 1 gggctaatta tggggtgtcg cccttattcg actctatagt gaagttccta ttctctagaa 61 agtataggaa cttctgaagt ggggcgggat gggtgaggtg gagtacgcgc ccggggagcc 121 caagggcacg ccctggcacc cgcaccgcgg cttcgagtac aagccgaagg cgcgcatgat 181 gaaataacat aaggtggtcc cgtcgaaagc cgaagcttac cgaagtatac acttaaattc 241 agtgcacgtt tgcttgttga gaggaaaggt tgtgtgcgga cgaatttttt tttgaaaaca 301 ttaaccctta cgtaacgcga gatctgaagt tcctattctc tagaaagtat aggaacttct 361 agggataaca gggtaataga tctggatcct aatgatgaca tgcgatattt gccgacttaa 421 aaagctcaag tgctccaaag aaaaaccgaa gtgcgccaag tgtctgaaga acaactggga 481 gtgtcgctac tctcccaaaa ccaaaaggtc tccgctgact agggcacatc tgacagaagt 541 ggaatcaagg ctagaaagac tggaacagct atttctactg atttttcctc gagaagacct 601 tgacatgatt ttgaaaatgg attctttaca ggatataaaa gcattgttaa caggattatt 661 tgtacaagat aatgtgaata aagatgccgt cacagataga ttggcttcag tggagactga 721 tatgcctcta acattgagac agcatagaat aagtgcgaca tcatcatcgg aagagagtag 781 taacaaaggt caaagacagt tgactgtatc gattgactcg gcagctcatc atgataactc 841 cacaattccg ttggatttta tgcccaggga tgctcttcat ggatttgatt ggtctgaaga 901 ggatgacatg tcggatggct tgcccttcct gaaaacggac cccaacaata atgggttctt 961 tggcgacggt tctctcttat gtattcttcg atctattggc tttaaaccgg aaaattacac 1021 gaactctaac gttaacaggc tcccgaccat gattacggat agatacacgt tggcttctag 1081 atccacaaca tcccgtttac ttcaaagtta tctcaataat tttcacccct actgccctat 1141 cgtgcactca ccgacgctaa tgatgttgta taataaccag attgaaatcg cgtcgaagga 1201 tcaatggcaa atccttttta actgcatatt agccattgga gcctggtgta tagaggggga 1261 atctactgat atagatgttt tttactatca aaatgctaaa tctcatttga cgagcaaggt 1321 cttcgagtca ggttccataa ttttggtgac agccctacat cttctgtcgc gatatacaca 1381 gtggaggcag aaaacaaata ctagctataa ttttcacagc ttttccataa gaatggccat 1441 atcattgggc ttgaataggg acctcccctc gtccttcagt gatagcagca ttctggaaca 1501 aagacgccga atttggtggt ctgtctactc ttgggagatc caattgtccc tgcttggatc 1561 ctcgcgtgag ggccgcggca gcctgctgac ctgcggcgat gtggaggaga accccgggcc 1621 ccctaggatg gataaagcgg aattaattcc cgagcctcca aaaaagaaga gaaaggtcga 1681 attgggatcc acgccgatgg agttccagta cctgcccgat acggatgacc gtcaccgtat 1741 cgaagaaaag cggaagcgaa cctatgaaac cttcaagtcc atcatgaaaa agtccccctt 1801 ctcgggcccc acggacccgc gccccccgcc ccgtcgtatt gcggttcctt cgcgcagcag 1861 tgccagcgtc cccaaacccg caccgcagcc ctacccgttc acttcctccc ttagcacgat 1921 taactatgat gagttcccca cgatggtgtt ccccagtgga caaatttccc aggcatcggc 1981 actggctccg gccccaccgc aagtcctccc ccaggcgccc gctccggcac cggctcccgc 2041 aatggtgagt gctctggccc aggcccccgc tccagtcccc gtgctggcgc ctggaccccc 2101 acaggcagtt gcccctcctg ctccgaaacc aacgcaggcg ggcgaaggaa ccctgagcga 2161 ggccctcttg cagcttcagt tcgatgacga agacttggga gccctgctgg gtaacagcac 2221 agaccctgcc gtattcaccg atctcgcatc cgtggacaac agcgagtttc agcagctctt 2281 gaatcaggga atcccggtcg cacctcatac cacagagccc atgctgatgg aatacccgga 2341 ggctatcacg cgactggtga ccggcgcaca gcgaccaccc gatccagccc ctgccccact 2401 gggtgccccg ggtttgccca atggcctcct cagcggcgat gaggatttct ccagcatcgc 2461 tgatatggat ttctccgctt tgctgagcca gataagctcc gctagcggag gaggtggtgg 2521 aggtggaggt ggaggtacta gtctggagat cgaggccgcc ttcctggagc gcgagaacac 2581 cgccctggag acccgcgtgg ccgagctgcg ccagcgcgtg cagcgcctgc gcaaccgcgt 2641 gagccagtac cgcacccgct acggccccct gggcggcggc aagtaaaagc ttatcgatac 2701 cgtcgactaa agccaaatag aaaattattc agttcctggc ttaagttttt aaaagtgata 2761 ttatttattt ggttgtaacc aaccaaaaga atgtaaataa ctaatacata attatgttag 2821 ttttaagtta gcaacaaatt gattttagct atattagcta cttggttaat aaatagaata 2881 tatttattta aagataattg cgtttttatt gtcagggagt gagtttgctt aaaaactcgt 2941 ttagatcgcg gccgcataac ttcgtatagc atacattata cgaagttatc atatgcccgg 3001 atgatcctga cgacggagac cgccgtcgtc gacaagccgg ccgagcatgc taagatacat 3061 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat 3121 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaccggttt 3181 acttgtacag ctcgtccatg ccgagagtga tcccggcggc ggtcacgaac tccagcagga 3241 ccatgtgatc gcgcttctcg ttggggtctt tgctcagggc ggactgggtg ctcaggtagt 3301 ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt gttctgctgg tagtggtcgg 3361 cgagctgcac gctgccgtcc tcgatgttgt ggcggatctt gaagtgggcc ttgatgccgt 3421 tcttctgctt gtcggcggtg atatagacgt tgtggctgat gtagttgtac tccagcttgt 3481 gccccaggat gttgccgtcc tccttgaagt cgatgccctt cagctcgatg cggttcacca 3541 gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta gttgccgtcg tccttgaaga 3601 agatggtacg ctcctggacg tagccttcgg gcatggcgga cttgaagaag tcgtgctgct 3661 tcatgtggtc ggggtagcgg ctgaagcact gcacgcccca ggtcagggtg gtcacgaggg 3721 tgggccaggg cacgggcagc ttgccggtgg tgcagatgaa cttcagggtc agcttgccgt 3781 aggtggcatc gccctcgccc tcgccggaca cgctgaacct gtggccgttt acgtcgccgt 3841 ccagctcgac caggatgggc accaccccgg tgaacagctc ctcgcccttg ctcaccatgt 3901 tgacgcgtcc gattgtttag cttgttcagc tgcgcttgtt tatttgctta gctttcgctt 3961 agcgacgtgt tcactttgct tgtttgaatt gaattgtcgc tccgtagacg aagcgcctct 4021 atttatactc cggcgtacgt agagtctccg ctcgcctgat tattttaatc tccgcctgat 4081 tattttaatg accgcctgat tattttaatc tacgcctgat tattttaaat gcatcgcctg 4141 attattttaa tctccgcctg attattttaa tgaccgcctg attattttaa tctacgcctg 4201 attattttaa cgggtcacgg aagttagacg tcataacttc gtatagcata cattatacga 4261 agttatagat cactagtcat ggcatcattg aaacagcaag gctcttacaa gttttcacaa 4321 aaatctatga actagacaaa acagtaactg cagaaaaaag tcctatatgt gcaaaaaaat 4381 gcttgatgat ttgtaatgag attgaggagg tttcgagaca ggcaccaaag tttttacaaa 4441 tggatatttc caccaccgct ctaaccaatt tgttgaagga acacccttgg ctatccttta 4501 caagattcga actgaagtgg aaacagttgt ctcttatcat ttatgtatta agagattttt 4561 tcactaattt tacccagaaa aagtcacaac tagaacagga tcaaaatgat catcaaagtt 4621 atgaagttaa acgatgctcc atcatgttaa gcgatgcagc acaaagaact gttatgtctg 4681 taagtagcta tatggacaat cataatgtca ccccatattt tgcctggaat tgttcttatt 4741 acttgttcaa tgcagtccta gtacccataa agactctact ctcaaactca aaatcgaatg 4801 ctgagaataa cgagaccgca caattattac aacaaattaa cactgttctg atgctattaa 4861 aaaaactggc cacttttaaa atccagactt gtgaaaaata cattcaagta ctggaagagg 4921 tatgtgcgcc gtttctgtta tcacagtgtg caatcccatt accgcatatc agttataaca 4981 atagtaatgg tagcgccatt aaaaatattg tcggttctgc aactatcgcc caatacccta 5041 ctcttccgga ggaaaatgtc aacaatatca gtgttaaata tgtttctcct ggctcagtag 5101 ggccttcacc tgtgccattg aaatcaggag caagtttcag tgatctagtc aagctgttat 5161 ctaaccgtcc accctctcgt aactctccag tgacaatacc aagaagcaca ccttcgcatc 5221 gctcagtcac gccttttcta gggcaacagc aacagctgca atcattagtg ccactgaccc 5281 cgtctgcttt gtttggtggc gccaatttta atcaaagtgg gaatattgct gatagctcat 5341 tgtccttcac tttcactaac agtagcaacg gtccgaacct cataacaact caaacaaatt 5401 ctcaagcgct ttcacaacca attgcctcct ctaacgttca tgataacttc atgaataatg 5461 aaatcacggc tagtaaaatt gatgatggta ataattcaaa accactgtca cctggttgga 5521 cggaccaaac tgcgtataac gcgtttggaa tcactacagg gatgtttaat accactacaa 5581 tggatgatgt atataactat ctattcgatg atgaagatac cccaccaaac ccaaaaaaag 5641 agtaatagac tagtgaagtt cctattctct agaaagtata ggaacttcta gtgatctgaa 5701 ttcattttca acgtcctcga tagtatagtg gttagtatcc ccgcctgtca cgcgggagac 5761 cggggttcaa ttccccgtcg gggagaatct gtgattcttt ttttttttct tttactttgt 5821 tatataaaca atttttgttt taattgaatc taatttgcca ttgcttttag gaatctcagg 5881 catccagcaa gcgtttgtcc gccgaatcgc ccatcagtga agaagatcct gtggcggcta 5941 cgaaaatctc cccggccatg tcggcctcca cctccagcga aaaacccatc agcgagctgg 6001 ccacctctgt gctgacccac cgctttccag actccacctc ctcacccggc gaacatggcc 6061 ttggacgaat gcagttgtcg atccgctaca gcgcccagcg tcaaaaacta gacgtgacca 6121 tacacaaaat ccagaagata ccacttcgcg atcccagcaa tatccccgat ccgtatgtta 6181 agctgtatct gttgcctgga cgcaccaagg agtcgaaacg caagacgagc gtgatcaagg 6241 acaactgcaa ccccgtctac gatgcatcct ttgagtacct gatttccatt gccgaactca 6301 ggcagacgga actggaggtg acggtgtgca cccaaaaggg attcctatcc ggcggtagtc 6361 ccatcattgg catggtaggt acccgaaagc aaccccttag ttacagacac agcgcgtacg 6421 tccttcgcat ccttatgatt cccaagtaca tattctgcaa gagtacagta tatataggaa 6481 agatatccgg gtgaacttcg atgtgcagcg taccacaacg ttttagagct agaaatagca 6541 agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt 6601 tgctcacctg tgattgctcc tactcaaata caaaaacatc aaattttctg tcaataaagc 6661 atatttattt atatttattt tacaggaaag aattcctttt aaagtgtatt ttaacctata 6721 atgaaaaacg attaaaaaaa atacataaaa taattcgaaa atttttgaat agcccaggtt 6781 gataaaaatt catttcatac gttttataac ttatgcccct aagtattttt tgaccatagt 6841 gtttcaattc tacattaatt ttacagagta gaatgaaacg ccacctactc agccaagagg 6901 cgaaaaggtt agctcgccaa gcagagaggg cgccagtgct cactactttt tataattctc 6961 aacttctttt tccagactca gttcgtatat atagacctat tttcaattta acgtcgtgta 7021 ttctgagaaa gctggagttt tagagctaga aatagcaagt taaaataagg ctagtccgtt 7081 atcaacttga aaaagtggca ccgagtcggt gcttttttgc ctacctggag cctgagagtt 7141 gttcaataaa ataaaaatgt ttcgtttttt tgctttcgcc agtatttatt atttttcatc 7201 aatatgtatt caatttggta tgtatttagt aattgtaata tatagacaat ggttttccgt 7261 tgacgtacat acatctgacg tgtgtttatt tagacataat agttatgttt tcacatcttt 7321 ttaatgttcg cttaatgcgt atgcttaatt aattctgacg taagctagtt acaaacccca 7381 cggacatgct aagggttaat caacaatcat atcgctgtct cactcagact caatacgaca 7441 ctcagaatac tattcctttc actcgcactt attgcaagca tacgttaagt ggatgtctct 7501 tgccgacggg accaccttat gttatttcat catgaaaatg aagtgaagtt cctatacttt 7561 ctagagaata ggaacttcta tagtgagtcg aataagggcg acacaaaatt tattctaaat 7621 gcataataaa tactgataac atcttatagt ttgtattata ttttgtatta tcgttgacat 7681 gtataatttt gatatcaaaa actgattttc cctttattat tttcgagatt tattttctta 7741 attctcttta acaaactaga aatattgtat atacaaaaaa tcataaataa tagatgaata 7801 gtttaattat aggtgttcat caatcgaaaa agcaacgtat cttatttaaa gtgcgttgct 7861 tttttctcat ttataaggtt aaataattct catatatcaa gcaaagtgac aggcgccctt 7921 aaatattctg acaaatgctc tttccctaaa ctccccccat aaaaaaaccc gccgaagcgg 7981 gtttttacgt tatttgcgga ttaacgatta ctcgttatca gaaccgccca gggggcccga 8041 gcttaagact ggccgtcgtt ttacaacaca gaaagagttt gtagaaacgc aaaaaggcca 8101 tccgtcaggg gccttctgct tagtttgatg cctggcagtt ccctactctc gccttccgct 8161 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac 8221 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga 8281 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat 8341 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac 8401 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct 8461 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg 8521 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg 8581 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt 8641 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg 8701 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg ggctaactac 8761 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga 8821 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt 8881 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt 8941 tctacggggt ctgacgctca gtggaacgac gcgcgcgtaa ctcacgttaa gggattttgg 9001 tcatgagctt gcgccgtccc gtcaagtcag cgtaatgctc tgcttaccaa tgcttaatca 9061 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg 9121 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagcgct gcgatgatac 9181 cgcgagaacc acgctcaccg gctccggatt tatcagcaat aaaccagcca gccggaaggg 9241 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc 9301 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccatcgcta 9361 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac 9421 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc 9481 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac 9541 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact 9601 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa 9661 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt 9721 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca 9781 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa 9841 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac 9901 tcatattctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg 9961 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggtcagtgtt acaaccaatt 10021 aaccaattct gaacattatc gcgagcccat ttatacctga atatggctca taacacccct 10081 tgtttgcctg gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg 10141 aaacgccgta gcgccgatgg tagtgtgggg actccccatg cgagagtagg gaactgccag 10201 gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgccc //