LOCUS       sequence-443876-       10763 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This vector enhances CRISPRi sgRNA cloning efficiency via Golden 
            Gate assembly, replacing the ccdB toxin gene for positive selection 
            of successful insertions..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 7549 to 7583)
  AUTHORS   Zhang Y, Zhang T, Xiao X, Wang Y, Kawalek A, Ou J, Ren A, Sun W, de 
            Bakker V, Liu Y, Li Y, Yang L, Ye L, Jia N, Veening JW, Liu X
  TITLE     CRISPRi screen identifies FprB as a synergistic target for gallium 
            therapy in Pseudomonas aeruginosa.
  JOURNAL   Nat Commun. 2025 Jul 1;16(1):5870. doi: 10.1038/s41467-025-61208-z.
  PUBMED    40595632
REFERENCE   2  (bases 1 to 10763)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2025-07-1"; volume: "16(1):5870. doi"; pages: " 
            10.1038/s41467-025-61208-z"
COMMENT     Sequence Label: pCRISPRi-ccdB
FEATURES             Location/Qualifiers
     source          1..10763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(423..442)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     542..564
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(602..620)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        688..792
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(793..861,862..1653)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1011..1030)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1954..2342
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            2611..2719
                     /note="incP origin of transfer"
     mobile_element  2823..2988
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     CDS             complement(3143..3232)
                     /codon_start=1
                     /product="3 tandem Myc epitope tags"
                     /label=3xMyc
                     /translation="EQKLISEEDLEQKLISEEDLEQKLISEEDL"
     CDS             complement(join(3242..4825,4826..4828,4829..7315,7316..7318,
                     7319..7345))
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     protein_bind    7392..7410
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7417..7435
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        7549..7583
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     misc_feature    8415..8433
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
     CDS             8515..8820
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     8699..8718
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     misc_feature    8921..8939
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
     protein_bind    complement(9228..9275)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        join(9352..9357,9358..9374,9375..9380)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             9491..10291
                     /codon_start=1
                     /gene="aac(3)IV"
                     /product="aminoglycoside 3-N-acetyltransferase type IV     
                               "
                     /label=ApmR
                     /note="confers resistance to apramycin (Paget and Davies, 
                     1996)"
                     /translation="MSSAVECNVVQYEWRKAELIGQLLNLGVTPGGVLLVHSSFRSVRP
                     LEDGPLGLIEALRAALGPGGTLVMPSWSGLDDEPFDPATSPVTPDLGVVSDTFWRLPNV
                     KRSAHPFAFAAAGPQAEQIISDPLPLPPHSPASPVARVHELDGQVLLLGVGHDANTTLH
                     LAELMAKVPYGVPRHCTILQDGKLVRVDYLENDHCCERFALADRWLKEKSLQKEGPVGH
                     AFARLIRSRDIVATALGQLGRDPLIFLHPPEAGCEECDAARQSIG"
     protein_bind    complement(10346..10393)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     terminator      complement(10444..10530)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(10633..10727)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(join(10755..10763,1..8))
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(join(10756..10763,1..9))
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
ORIGIN
        1 cgacctcgaa ccccacgccc ctctttaata cgacgggcaa tttgcacttc agaaaatgaa
       61 gagtttgctt tagccataac aaaagtccag tatgcttttt cacagcataa ctggactgat
      121 ttcagtttac aactattctg tctagtttaa gactttattg tcatagttta gatctatttt
      181 gttcagttta agactttatt gtccgcccac acccgcttac gcagggcatc catttattac
      241 tcaaccgtaa ccgattttgc caggttacgc ggctggtcct ctagctggcg taatagcgaa
      301 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg
      361 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc
      421 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
      481 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
      541 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
      601 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
      661 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
      721 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
      781 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
      841 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
      901 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
      961 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     1021 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     1081 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     1141 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     1201 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     1261 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     1321 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     1381 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     1441 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     1501 gagcgtgggt cccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     1561 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     1621 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     1681 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     1741 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     1801 gtagaaaaga tcaaaggcag ttattggtgc ctcactgatt aagcattggt aactgtcaga
     1861 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttttgcggcc
     1921 gcaagatccg gccacgatgc gtccggcgta gaggatctga agatcagcag ttcaacctgt
     1981 tgatagtacg tactaagctc tcatgtttca cgtactaagc tctcatgttt aacgtactaa
     2041 gctctcatgt ttaacgaact aaaccctcat ggctaacgta ctaagctctc atggctaacg
     2101 tactaagctc tcatgtttca cgtactaagc tctcatgttt gaacaataaa attaatataa
     2161 atcagcaact taaatagcct ctaaggtttt aagttttata agaaaaaaaa gaatatataa
     2221 ggcttttaaa gcttttaagg tttaacggtt gtggacaaca agccagggat gtaacgcact
     2281 gagaagccct tagagcctct caaagcaatt ttgagtgaca caggaacact taacggctga
     2341 catgggaatt agccatggca tcacagtatc gtgatgacag aggcagggag tgcggccttt
     2401 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcaggg
     2461 gattccttaa ggtatacttt ccgctgcata accctgcttc ggggtcatta tagcgatttt
     2521 ttcggtatat ccatcctttt tcgcacgata tacaggattt tgccaaaggg ttcgtgtaga
     2581 ctttccttgg tgtatccaac ggcgtcagcc gggcaggata ggtgaagtag gcccacccgc
     2641 gagcgggtgt tccttcttca ctgtccctta ttcgcacctg gcggtgctca acgggaatcc
     2701 tgctctgcga ggctggccga taagctctga taccgctcgc cgcagccgaa cgaccgagcg
     2761 cagcgggtca gtgagcgagg aagcggaaga gcccaaacat aacaggaaga aaaatgcccc
     2821 gctgtgggcg gacaaaatag ttgggaactg ggaggggtgg aaatggagtt tttaaggatt
     2881 atttagggaa gagtgacaaa atagatggga actgggtgta gcgtcgtaag ctaatacgaa
     2941 aattaaaaat gacaaaatag tttggaacta gatttcactt atctggttct tgaggcggtt
     3001 aaaagagccg tactcttctc cgatgtcgac taggccatga tgctcattct gtgggaccaa
     3061 aacgaaaaaa caccctttcg ggtgtctttt ctggaatttg gtaccgaggc tgcaagtccc
     3121 attaaggagg cgcgccttat tacagatctt cctcagagat gagcttctgt tccagatctt
     3181 cttctgaaat caacttttgt tccagatctt cttcagagat gagtttctgc tccgcggccg
     3241 cgtcacctcc tagctgactc aaatcaatgc gtgtttcata aagaccagtg atggattgat
     3301 ggataagagt ggcatctaaa acttcttttg tagacgtata tcgtttacga tcaattgttg
     3361 tatcaaaata tttaaaagca gcgggagctc caagattcgt caacgtaaat aaatgaataa
     3421 tattttctgc ttgttcacgt attggtttgt ctctatgttt gttatatgca ctaagaactt
     3481 tatctaaatt ggcatctgct aaaataacac gcttagaaaa ttcactgatt tgctcaataa
     3541 tctcatctaa ataatgctta tgctgctcca caaacaattg tttttgttcg ttatcttctg
     3601 gactaccctt caacttttca taatgactag ctaaatataa aaaattcaca tatttgcttg
     3661 gcagagccag ctcatttcct ttttgtaatt ctccggcact agccagcatc cgtttacgac
     3721 cgttttctaa ctcaaaaaga ctatatttag gtagtttaat gattaagtct tttttaactt
     3781 ccttatatcc tttagcttct aaaaagtcaa tcggattttt ttcaaaggaa cttctttcca
     3841 taattgtgat ccctagtaac tctttaacgg attttaactt cttcgatttc cctttttcca
     3901 ccttagcaac cactaggact gaataagcta ccgttggact atcaaaacca ccatattttt
     3961 ttggatccca gtctttttta cgagcaataa gcttgtccga atttcttttt ggtaaaattg
     4021 actccttgga gaatccgcct gtctgtactt ctgttttctt gacaatattg acttggggca
     4081 tggacaatac tttgcgcact gtggcaaaat ctcgcccttt atcccagaca atttctccag
     4141 tttccccatt agtttcgatt agagggcgtt tgcgaatctc tccatttgca agtgtaattt
     4201 ctgttttgaa gaagttcatg atattagagt aaaagaaata ttttgcggtt gctttgccta
     4261 tttcttgctc agacttagca atcattttac gaacatcata aactttataa tcaccataga
     4321 caaactccga ttcaagtttt ggatatttct taatcaaagc agttccaacg acggcattta
     4381 gatacgcatc atgggcatga tggtaattgt taatctcacg tactttatag aattggaaat
     4441 cttttcggaa gtcagaaact aatttagatt ttaaggtaat cactttaacc tctcgaataa
     4501 gtttatcatt ttcatcgtat ttagtattca tgcgactatc caaaatttgt gccacatgct
     4561 tagtgatttg gcgagtttca accaattggc gtttgataaa accagcttta tcaagttcac
     4621 tcaaacctcc acgttcagct ttcgttaaat tatcaaactt acgttgagtg attaacttgg
     4681 cgtttagaag ttgtctccaa tagtttttca tctttttgac tacttcttca cttggaacgt
     4741 tatccgattt accacgattt ttatcagaac gcgttaagac cttattgtct attgaatcgt
     4801 ctttaaggaa actttgtgga acaatggcat cgacatcata atcacttaaa cgattaatat
     4861 ctaattcttg gtccacatac atgtctcttc cattttggag ataatagaga tagagctttt
     4921 cattttgcaa ttgagtattt tcaacaggat gctctttaag aatctgactt cctaattctt
     4981 tgataccttc ttcgattcgt ttcatacgct ctcgcgaatt tttctggccc ttttgagttg
     5041 tctgattttc acgtgccatt tcaataacga tattttctgg cttatgccgc cccattactt
     5101 tgaccaattc atcaacaact tttacagtct gtaaaatacc ttttttaata gcagggctac
     5161 cagctaaatt tgcaatatgt tcatgtaaac tatcgccttg tccagacact tgtgcttttt
     5221 gaatatcttc tttaaatgtc aaactatcat catggatcag ctgcataaaa ttgcgattgg
     5281 caaaaccatc tgatttcaaa aaatctaata ttgttttgcc agattgctta tccctaatac
     5341 cattaatcaa ttttcgagac aaacgtcccc aaccagtata acggcgacgt ttaagctgtt
     5401 tcatcacctt atcatcaaag aggtgagcat atgttttaag tctttcctca atcatctccc
     5461 tatcttcaaa taaggtcaat gttaaaacaa tatcctctaa gatatcttca ttttcttcat
     5521 tatccaaaaa atctttatct ttaataattt ttagcaaatc atggtaggta cctaatgaag
     5581 cattaaatct atcttcaact cctgaaattt caacactatc aaaacattct atttttttga
     5641 aataatcttc ttttaattgc ttaacggtta cttttcgatt tgttttgaag agtaaatcaa
     5701 caatggcttt cttctgttca cctgaaagaa atgctggttt tcgcattcct tcagtaacat
     5761 atttgacctt tgtcaattcg ttataaaccg taaaatactc ataaagcaaa ctatgttttg
     5821 gtagtacttt ttcatttgga agatttttat caaagtttgt catgcgttca ataaatgatt
     5881 gagctgaagc acctttatcg acaacttctt caaaattcca tggggtaatt gtttcttcag
     5941 acttccgagt catccatgca aaacgactat tgccacgcgc caatggacca acataataag
     6001 ggattcgaaa agtcaagatt ttttcaatct tctcacgatt gtcttttaaa aatggataaa
     6061 aatcttcttg tcgtctcaaa atagcatgca gctcacccaa gtgaatttga tggggaatag
     6121 agccgttgtc aaaggtccgt tgcttgcgca gcaaatcttc acgatttagt ttcaccaata
     6181 attcctcagt accatccatt ttttctaaaa ttggtttgat aaatttataa aattcttctt
     6241 ggctagctcc cccatcaata taacctgcat atccgttttt tgattgatca aaaaagattt
     6301 ctttatactt ttctggaagt tgttgtcgaa ctaaagcttt taaaagagtc aagtcttgat
     6361 gatgttcatc gtagcgttta atcattgaag ctgatagggg agccttagtt atttcagtat
     6421 ttactcttag gatatctgaa agtaaaatag catctgataa attcttagct gccaaaaaca
     6481 aatcagcata ttgatctcca atttgcgcca ataaattatc taaatcatca tcgtaagtat
     6541 cttttgaaag ctgtaattta gcatcttctg ccaaatcaaa atttgattta aaattagggg
     6601 tcaaacccaa tgacaaagca atgagattcc caaataagcc atttttcttc tcaccgggga
     6661 gctgagcaat gagattttct aatcgtcttg atttactcaa tcgtgcagaa agaatcgctt
     6721 tagcatctac tccacttgcg ttaatagggt tttcttcaaa taattgattg taggtttgta
     6781 ccaactggat aaatagtttg tccacatcac tattatcagg atttaaatct ccctcaatca
     6841 aaaaatgacc acgaaactta atcatatgcg ctaaggccaa atagattaag cgcaaatccg
     6901 ctttatcagt agaatctacc aatttttttc gcagatgata gatagttgga tatttctcat
     6961 gataagcaac ttcatctact atatttccaa aaataggatg acgttcatgc ttcttatctt
     7021 cttccaccaa aaaagactct tcaagtcgat gaaagaaact atcatctact ttcgccatct
     7081 catttgaaaa aatctcctgt agataacaaa tacgattctt ccgacgtgta taccttctac
     7141 gagctgtccg tttgagacga gtcgcttccg ctgtctctcc actgtcaaat aaaagagccc
     7201 ctataagatt ttttttgata ctgtggcggt ctgtatttcc cagaaccttg aactttttag
     7261 acggaacctt atattcatca gtgatcaccg cccatccgac gctatttgtg ccgatagcta
     7321 agcctattga gtatttctta tccatactag ttttcctgtg tgaaacctgc tgatgtgctc
     7381 tgtgctcagt atctctatca ctgataggga tgtcaatctc tatcactgat agggaactag
     7441 tgacgtgttg atcaccgtct agatccgcga aaaaaccccg ccgaagcggg gttttttgcg
     7501 gaattcgagc tcggtacctc gcgaatgcat ctagatatcg gatcccaatt gacggctagc
     7561 tcagtcctag gtacagtgct agctcacaca ggaaaggcct cgatgagccg gctcgataaa
     7621 agcaaggtga tcaacagcgc gctggaactc ctcaatgaag tcggcatcga gggcctgacc
     7681 acccgcaaac tcgcccagaa gctcggggtc gagcagccca ccctgtattg gcatgtcaaa
     7741 aataaacgcg ccctcctgga tgccctggcc atcgagatgc tcgaccgcca tcacacgcac
     7801 ttctgcccgc tcgaggggga atcgtggcag gatttcctcc ggaataatgc gaagtccttc
     7861 cgctgcgcgc tcctctccca tcgcgatggg gccaaggtcc atctcggcac ccgcccgacc
     7921 gagaagcaat atgagacgct cgaaaaccaa ctggcgttcc tctgccaaca ggggttcagc
     7981 ctcgaaaatg cgctgtatgc gctctccgcc gtggggcatt tcaccctggg ctgcgtgctg
     8041 gaggaccaag agcatcaggt cgccaaagag gaacgcgaaa cgccgaccac ggattccatg
     8101 cccccgctcc tccgccaagc gatcgagctc ttcgatcatc agggcgcgga gccggccttc
     8161 ctcttcggcc tcgagctcat catctgcggc ctcgagaagc agctgaaatg cgaatccggc
     8221 tcctaaggat ctaaatagga ggaaaattac gcaaaaaacc ccgcttcggc ggggtttttt
     8281 cgcgagctcc actgtaacag agcattagcg caaggtgatt tttgtcttct tgcgctaatt
     8341 tttttgcgac tactcttgcc tactacctat cgactgagct gaaagaattc ctaaagatct
     8401 tcgtcggcag cgtcagatgt gtataagaga cagccattct acagtttatt cttgacattg
     8461 cactgtcccc ctggtataat aactatatga gacccgagaa caggggctgg tgaaatgcag
     8521 tttaaggttt acacctataa aagagagagc cgttatcgtc tgtttgtgga tgtacagagt
     8581 gatattattg acacgcccgg gcgacggatg gtgatccccc tggccagtgc acgtctgctg
     8641 tcagataaag tctcccgtga actttacccg gtggtgcata tcggggatga aagctggcgc
     8701 atgatgacca ccgatatggc cagtgtgccg gtttccgtta tcggggaaga agtggctgat
     8761 ctcagccacc gcgaaaatga catcaaaaac gccattaacc tgatgttctg gggaatataa
     8821 ggtctcagtt taagagctat gctggaaaca gcatagcaag tttaaataag gctagtccgt
     8881 tatcaacttg aaaaagtggc accgagtcgg tgcttttttt ctgtctctta tacacatctc
     8941 cgagcccacg agacattcat gtggctgacc gttctgttgt ctctcgctct tccgagtaga
     9001 cgaacaataa ggcctcccta acggggggcc ttttttattg ataacaaaag tcagtgcttc
     9061 cgctatttcc aaaataccgg gctaatacgg tttaaacgac ctcctggatt tgctcagaca
     9121 gccttttcgt cattcgtttc agccaaaaaa cttaagaccg ccggtcttgt ccactacctt
     9181 gcagtaatgc ggtggacagg atcggcggtt ttcttttctc ttctcaagaa gttcctatac
     9241 tttctagaga ataggaactt cggaatagga acttcctcct gaacggccat aagaacgaag
     9301 gctgtctgtt gaactctcga gccgtggaaa cggatgaagg cacgaaccca gttgacataa
     9361 gcctgttcgg ttcgtaaact gtaatgcaag tagcgtatgc gctcaaaact ggatggcttt
     9421 cttgccgcca aggatctgat ggcgcagggg atcaagatct gatcaagaga caggatgagg
     9481 atcgtttcgc atgtcatcag cggtggagtg caatgtcgtg caatacgaat ggcgaaaagc
     9541 cgagctcatc ggtcagcttc tcaaccttgg ggttaccccc ggcggtgtgc tgctggtcca
     9601 cagctccttc cgtagcgtcc ggcccctcga agatgggcca cttggactga tcgaggccct
     9661 gcgtgctgcg ctgggtccgg gagggacgct cgtcatgccc tcgtggtcag gtctggacga
     9721 cgagccgttc gatcctgcca cgtcgcccgt tacaccggac cttggagttg tctctgacac
     9781 attctggcgc ctgccaaatg taaagcgcag cgcccatcca tttgcctttg cggcagcggg
     9841 gccacaggca gagcagatca tctctgatcc attgcccctg ccacctcact cgcctgcaag
     9901 cccggtcgcc cgtgtccatg aactcgatgg gcaggtactt ctcctcggcg tgggacacga
     9961 tgccaacacg acgctgcatc ttgccgagtt gatggcaaag gttccctatg gggtgccgag
    10021 acactgcacc attcttcagg atggcaagtt ggtacgcgtc gattatctcg agaatgacca
    10081 ctgctgtgag cgctttgcct tggcggacag gtggctcaag gagaagagcc ttcagaagga
    10141 aggtccagtc ggtcatgcct ttgctcggtt gatccgctcc cgcgacattg tggcgacagc
    10201 cctgggtcaa ctgggccgag atccgttgat cttcctgcat ccgccagagg cgggatgcga
    10261 agaatgcgat gccgctcgcc agtcgattgg ctgagcggga ctctggggtt cgaaatgact
    10321 cgagcatcag cttcaaaagc gctctgaagt tcctatactt tctagagaat aggaacttcg
    10381 gaataggtac ttcaagatcc ccaattcgag ctcatcgtcc gggccgcaag ctcctagcgg
    10441 cggatttgtc ctactcagga gagcgttcac cgacaaacaa cagataaaac gaaaggccca
    10501 gtctttcgac tgagcctttc gttttatttg atgcctcaag ctagagagtc attaccccag
    10561 gcgtttaagg gcaccaataa ctgccttaaa aaaattacgc cccgccctgc cactcatcgc
    10621 agtctagctt ggattctcac caataaaaaa cgcccggcgg caaccgagcg ttctgaacaa
    10681 atccagatgg agttctgagg tcattactgg atctatcaac aggagtccaa gctcagctaa
    10741 ttaagctagc ttatcgatac cgt
//
