LOCUS       sequence-307459-       15720 bp    DNA     circular SYN 14-APR-2025
DEFINITION  2lox landing pad plasmid with mariner transposon and transposase 
            used for CRAGE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15720)
  AUTHORS   Wang G, Zhao Z, Ke J, Engel Y, Shi YM, Robinson D, Bingol K, Zhang 
            Z, Bowen B, Louie K, Wang B, Evans R, Miyamoto Y, Cheng K, Kosina S,
            De Raad M, Silva L, Luhrs A, Lubbe A, Hoyt DW, Francavilla C, Otani 
            H, Deutsch S, Washton NM, Rubin EM, Mouncey NJ, Visel A, Northen T, 
            Cheng JF, Bode HB, Yoshikuni Y
  TITLE     CRAGE enables rapid activation of biosynthetic gene clusters in 
            undomesticated bacteria.
  JOURNAL   Nat Microbiol. 2019 Dec;4(12):2498-2510. doi: 
            10.1038/s41564-019-0573-8. Epub 2019 Oct 14.
  PUBMED    31611640
REFERENCE   2  (bases 1 to 15720)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41564-019-0573-8"; journalName: "Nat Microbiol"; date: 
            "2019-12"; volume: "4"; issue: "12"; pages: "2498-2510"
COMMENT     Sequence Label: pW17
FEATURES             Location/Qualifiers
     source          1..15720
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1244..1260
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1268..1274,1275..1292,1293..1298))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1313..1334
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     oriT            1481..1589
                     /note="incP origin of transfer"
     primer_bind     1634..1652
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1677..1693)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(1994..2027)
                     /label=lox5171
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTGTAC) (Shaw et al., 2021). lox5171 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             complement(2065..2859)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHTGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2176..2195)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2786..2805
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(3394..4425)
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     misc_feature    4634..4741
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     4650..4666
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     4651..4667
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4701..4717)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4701..4717)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    4767..4800
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4861..4938
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4939..6021
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4958..4977)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6070..6075,6076..6093,6094..6100)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    6108..6124
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6132..6148
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6132..6148
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             6144..6593
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGI"
     primer_bind     complement(6164..6180)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      10447..11061
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"
     rep_origin      10538..11069
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      11137..11356
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     CDS             11447..12202
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     misc_feature    12205..12455
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             12781..13956
                     /codon_start=1
                     /gene="sopA"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopA
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQTNHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     CDS             13956..14927
                     /codon_start=1
                     /gene="sopB"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopB
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
     misc_feature    15000..15473
                     /gene="sopC"
                     /label=sopC
                     /note="centromere-like partitioning region of the bacterial
                     F plasmid"
ORIGIN
        1 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
       61 atccatcaca ctggcggccg ctcgagcatg catctagtaa cggccgccag tgtgctggaa
      121 ttctgcagat ccggtctaac aaagaaaaac acattttttt gtgaaaattc gtttttatta
      181 ttcaacatag ttcccttcaa gagcgataca acgattataa cgaccttcca attttttgat
      241 accattttgg tagtactcct tcggttttgc ctcaaaatag gcctcagttt cggcgatcac
      301 ctcttcattg cagccaaatt ttttccctgc gagcatcctt ttgaggtctg agaacaagaa
      361 aaagtcgctg ggggccagat ctggagaata cggcgggtgg ggaagcaatt cgaagcccaa
      421 ttcatgaatt tttgccatcg ttctcaatga cttgtggcac ggtgcgttgt cttggtggaa
      481 caacactttt ttcttcttca tgtggggccg ttttgccgcg atttcgacct tcaaacgctc
      541 caataacgcc atataatagt cactgttgat ggtttttccc ttctcaagat aatcgataaa
      601 aattattcca tgcgcatccc aaaaaacaga ggccattact ttgccagcgg acttttgagt
      661 ctttccacgc ttcggagacg gttcaccggt cgctgtccac tcagccgact gtcgattgga
      721 ctcaggagtg tagtgatgga gccatgtttc atccattgtc acatatcgac ggaaaaactc
      781 gggtgtatta cgagttaaca gctgcaaaca ccgctcagaa tcatcaacac gttgttgttt
      841 ttggtcaaat gtgagctcgc gcggcaccca ttttgcacag agcttccgca tatccaaata
      901 ttgatgaatg atatgaccaa cacgttcctt tgatatcttt aaggcctctg ctatctcgat
      961 caacttcatt ttacggtcat tcaaaatcat tttgtggatt tttttgatgt tttcgtcggt
     1021 aaccacctct ttcgggcgtc cactgcgttc accgtcctcc gtgctcattt caccacgctt
     1081 gaattttgca taccaatcaa ttattgttga tttccctggg gcagagtccg gaaactcatt
     1141 atcaagccaa gtttttgctt ccactgtatt ttttcccttc agaaaacagt attttatcaa
     1201 aacacgaaat tccttttttt ccatatgtgt ttcctgtgtg aaattgttat ccgctcacaa
     1261 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     1321 gctaactcac attaattgcg ttgcgctcac ccgctgcata accctgcttc ggggtcatta
     1381 tagcgatttt ttcggtatat ccatcctttt tcgcacgata tacaggattt tgccaaaggg
     1441 ttcgtgtaga ctttccttgg tgtatccaac ggcgtcagcc gggcaggata ggtgaagtag
     1501 gcccacccgc gagcgggtgt tccttcttca ctgtccctta ttcgcacctg gcggtgctca
     1561 acgggaatcc tgctctgcga ggctggccga taagctctaa gaaaccatta ttatcatgac
     1621 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcaagaat taattcactg
     1681 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaaca cataacaggt
     1741 tggctgataa gtccccggtc tggccggccc attgcatcct gcagggcgta atgctctgcc
     1801 agtgtcgcga accccagagt cgtcgacacg cgcagaccaa aacgatctca agaagatcat
     1861 cttattaatc agataaaata tttctaggca ccaataactg ccttaaaaaa attacgcccc
     1921 gccctgccac tcatcgcagt actgttgata ccgggaagtc ctgggccaac ttttggcgaa
     1981 aatgagacgt tgaataactt cgtatagtac acattatacg aagttattcg gcacgtaaga
     2041 ggttccaact ttcacccggg ccgctcagaa gaactcgtca agaaggcgat agaaggcgat
     2101 gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc
     2161 gccaagctct tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac
     2221 acccagccgg ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg
     2281 caagcaggca tcgccatggg tcacgacgag atcctcgccg tcgggcatcc gcgccttgag
     2341 cctggcgaac agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc
     2401 gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc
     2461 gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga
     2521 tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa
     2581 tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc
     2641 cgtcgtggcc agccacgata gccgcgctgc ctcgtcttgg agttcattca gggcaccgga
     2701 caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc
     2761 atcagagcag ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc
     2821 ggccggagaa cctgtgtgca atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt
     2881 ctcttgatca gatcttgatc ccctgcgcca tcagatcctt ggcggcaaga aagccatcca
     2941 gtttactttg cagggcttcc caaccttacc agagggcgcc ccagctggca attccggttc
     3001 gcttgctgtc cataaaaccg cccagtctag ctatcgccat gtaagcccac tgcaagctac
     3061 ctgctttctc tttgcgcttg cgttttccct tgtccagata gcccagtagc tgacattcat
     3121 ccggggtcag caccgtttct gcggactggc tttctacgtg ttccgcttcc tttagcagcc
     3181 cttgcgccct gagtgcttgc ggcagcgtga actagttaag gttcagcagt tcctgccctc
     3241 tgattttcca gtctgaccac ttcggattat cccgtgacag gtcattcaga ctggctaatg
     3301 cacccagtaa ggcagcggta tcatcaacag gcttacccgt cttactgtct cgagttttgt
     3361 tcaaaaaaaa gcccgctcat taggcgggct gggctaatcg ccatcttcca gcaggcgcac
     3421 cattgcccct gtttcactat ccaggttacg gatatagttc atgacaatat ttacattggt
     3481 ccagccacca gcttgcatga tctccggtat tgaaactcca gcgcgggcca tatctcgcgc
     3541 ggctccgaca cgggcactgt gtccagacca ggccaggtat ctctgaccag agtcatcctt
     3601 agcgccgtaa atcaatcgat gagttgcttc aaaaatccct tccagggcgc gagttgatag
     3661 ctggctggtg gcagatggcg cggcaacacc attttttctg acccggcaaa acaggtagtt
     3721 attcggatca tcagctacac cagagacgga aatccatcgc tcgaccagtt tagttacccc
     3781 caggctaagt gccttctcta cacctgcggt gctaaccagc gttttcgttc tgccaatatg
     3841 gattaacatt ctcccaccgt cagtacgtga gatatcttta accctgatcc tggcaatttc
     3901 ggctatacgt aacagggtgt tataagcaat ccccagaaat gccagattac gtatatcctg
     3961 gcagcgatcg ctattttcca tgagtgaacg aacctggtcg aaatcagtgc gttcgaacgc
     4021 tagagcctgt tttgcacgtt caccggcatc aacgttttct tttcggatcc gccgcataac
     4081 cagtgaaaca gcattgctgt cacttggtcg tggcagcccg gaccgacgat gaagcatgtt
     4141 tagctggccc aaatgttgct ggatagtttt tactgccaga ccgcgcgcct gaagatatag
     4201 aagataatcg cgaacatctt caggttctgc gggaaaccat ttccggttat tcaacttgca
     4261 ccatgccgcc cacgaccggc aaacggacag aagcattttc caggtatgct cagaaaacgc
     4321 ctggcgatcc ctgaacatgt ccatcaggtt cttgcgaacc tcatcactcg ttgcatcgac
     4381 cggtaatgca ggcaaatttt ggtgtacggt cagtaaattg gacatttttg cctcctaaaa
     4441 taaaaagttt aaattaaatc cataatgagt ttgatgattt caataatagt tttaatgacc
     4501 tccgaaatta gtttaatatg ctttaatttt tctttttcaa aatatctctt caaaaaatat
     4561 tacccaatac ttaataataa atagattata acacaaaatt cttttaaaaa agtagtttat
     4621 tttgttatca ttcggtaccg ggccccccct cgaggtcgac ggtatcgata agcttgatat
     4681 cgaattcctg cagcccgggg gatccactag ttctagagcg gccgccaccg cggtggagct
     4741 cactcaaagg cggtaatacg gttatcataa cttcgtataa tgtatgctat acgaagttat
     4801 cctttgcact ggattgcgag gctttgcacg atgcgtccgg cgtagaggat ccagatcccg
     4861 gacaccatcg aatggcgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
     4921 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
     4981 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
     5041 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
     5101 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
     5161 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
     5221 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
     5281 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
     5341 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
     5401 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
     5461 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
     5521 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
     5581 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
     5641 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
     5701 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
     5761 gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc
     5821 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     5881 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     5941 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     6001 cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagctca ctcattaggc
     6061 accccaggct ttacacttta tgcttccggc tcgtataatg tgtggaattg tgagcggata
     6121 acaatttcac acaggaaaca gctatgacca tgattacgga ttcactggcc gtcgttttac
     6181 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     6241 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     6301 gcagcctgaa tggcgaatgg cgctttgcct ggtttccggc accagaagcg gtgccggaaa
     6361 gctggctgga gtgcgatctt cctgaggccg atactgtcgt cgtcccctca aactggcaga
     6421 tgcacggtta cgatgcgccc atctacacca acgtgaccta tcccattacg gtcaatccgc
     6481 cgtttgttcc cacggagaat ccgacgggtt gttactcgct cacatttaat gttgatgaaa
     6541 gctggctaca ggaaggccag acgcgaatta tttttgatgg cgtcgggatc tgatccggat
     6601 ttactaactg gaagaggcac taaatgaaca cgattaacat cgctaagaac gacttctctg
     6661 acatcgaact ggctgctatc ccgttcaaca ctctggctga ccattacggt gagcgtttag
     6721 ctcgcgaaca gttggccctt gagcatgagt cttacgagat gggtgaagca cgcttccgca
     6781 agatgtttga gcgtcaactt aaagctggtg aggttgcgga taacgctgcc gccaagcctc
     6841 tcatcactac cctactccct aagatgattg cacgcatcaa cgactggttt gaggaagtga
     6901 aagctaagcg cggcaagcgc ccgacagcct tccagttcct gcaagaaatc aagccggaag
     6961 ccgtagcgta catcaccatt aagaccactc tggcttgcct aaccagtgct gacaatacaa
     7021 ccgttcaggc tgtagcaagc gcaatcggtc gggccattga ggacgaggct cgcttcggtc
     7081 gtatccgtga ccttgaagct aagcacttca agaaaaacgt tgaggaacaa ctcaacaagc
     7141 gcgtagggca cgtctacaag aaagcattta tgcaagttgt cgaggctgac atgctctcta
     7201 agggtctact cggtggcgag gcgtggtctt cgtggcataa ggaagactct attcatgtag
     7261 gagtacgctg catcgagatg ctcattgagt caaccggaat ggttagctta caccgccaaa
     7321 atgctggcgt agtaggtcaa gactctgaga ctatcgaact cgcacctgaa tacgctgagg
     7381 ctatcgcaac ccgtgcaggt gcgctggctg gcatctctcc gatgttccaa ccttgcgtag
     7441 ttcctcctaa gccgtggact ggcattactg gtggtggcta ttgggctaac ggtcgtcgtc
     7501 ctctggcgct ggtgcgtact cacagtaaga aagcactgat gcgctacgaa gacgtttaca
     7561 tgcctgaggt gtacaaagcg attaacattg cgcaaaacac cgcatggaaa atcaacaaga
     7621 aagtcctagc ggtcgccaac gtaatcacca agtggaagca ttgtccggtc gaggacatcc
     7681 ctgcgattga gcgtgaagaa ctcccgatga aaccggaaga catcgacatg aatcctgagg
     7741 ctctcaccgc gtggaaacgt gctgccgctg ctgtgtaccg caaggacaag gctcgcaagt
     7801 ctcgccgtat cagccttgag ttcatgcttg agcaagccaa taagtttgct aaccataagg
     7861 ccatctggtt cccttacaac atggactggc gcggtcgtgt ttacgctgtg tcaatgttca
     7921 acccgcaagg taacgatatg accaaaggac tgcttacgct ggcgaaaggt aaaccaatcg
     7981 gtaaggaagg ttactactgg ctgaaaatcc acggtgcaaa ctgtgcgggt gtcgataagg
     8041 ttccgttccc agagcgcatc aagttcattg aggaaaacca cgagaacatc atggcttgcg
     8101 ctaagtctcc actggagaac acttggtggg ctgagcaaga ttctccgttc tgcttccttg
     8161 cgttctgctt tgagtacgct ggggtacagc accacggcct gagctataac tgctcccttc
     8221 cgctggcgtt tgacgggtct tgctctggca tccagcactt ctccgcgatg ctccgagatg
     8281 aggtaggtgg tcgcgcggtt aacttgcttc ctagtgaaac cgttcaggac atctacggga
     8341 ttgttgctaa gaaagtcaac gagattctac aagcagacgc aatcaatggg accgataacg
     8401 aagtagttac cgtgaccgat gagaacactg gtgaaatctc tgagaaagtc aagctgggca
     8461 ctaaggcact ggctggtcaa tggctggctt acggtgttac tcgcagtgtg actaagcgtt
     8521 cagtcatgac gctggcttac gggtccaaag agttcggctt ccgtcaacaa gtgctggaag
     8581 ataccattca gccagctatt gattccggca agggtctgat gttcactcag ccgaatcagg
     8641 ctgctggata catggctaag ctgatttggg aatctgtgag cgtgacggtg gtagctgcgg
     8701 ttgaagcaat gaactggctt aagtctgctg ctaagctgct ggctgctgag gtcaaagata
     8761 agaagactgg agagattctt cgcaagcgtt gcgctgtgca ttgggtaact cctgatggtt
     8821 tccctgtgtg gcaggaatac aagaagccta ttcagacgcg cttgaacctg atgttcctcg
     8881 gtcagttccg cttacagcct accattaaca ccaacaaaga tagcgagatt gatgcacaca
     8941 aacaggagtc tggtatcgct cctaactttg tacacagcca agacggtagc caccttcgta
     9001 agactgtagt gtgggcacac gagaagtacg gaatcgaatc ttttgcactg attcacgact
     9061 ccttcggtac cattccggct gacgctgcga acctgttcaa agcagtgcgc gaaactatgg
     9121 ttgacacata tgagtcttgt gatgtactgg ctgatttcta cgaccagttc gctgaccagt
     9181 tgcacgagtc tcaattggac aaaatgccag cacttccggc taaaggtaac ttgaacctcc
     9241 gtgacatctt agagtcggac ttcgcgttcg cgtaacgcca aatcaatacg actccggatc
     9301 cacaggacgg gtgtggtcgc gaccgagata gggttgagtg ttgttccagt ttgagatctg
     9361 ggaatcattt gaaggttggt actatataaa aataatatgc atttaatact agcgacgcca
     9421 tctatgtgtc agaccgggga cttatcagcc aacctgttag cagaacttta aaagtgctca
     9481 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     9541 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     9601 tttctgggtg agcaaaaaca ggaaggcaaa atgcacaggg acaccaggat ttatttattc
     9661 tgcgaagtga tcttccgtca caggtattta ttcgcgataa gctcatggag cggcgtaacc
     9721 gtcgcacagg aaggacagag aaagcgcgga tctgggaagt gacggacaga acggtcagga
     9781 cctggattgg ggaggcggtt gccgccgctg ctgctgacgg tgtgacgttc tctgttccgg
     9841 tcacaccaca tacgttccgc cattcctatg cgatgcacat gctgtatgcc ggtataccgc
     9901 tgaaagttct gcaaagcctg atgggacata agtccatcag ttcaacggaa gtctacacga
     9961 aggtttttgc gctggatgtg gctgcccggc accgggtgca gtttgcgatg ccggagtctg
    10021 atgcggttgc gatgctgaaa caattatcct gagaataaat gccttggcct ttatatggaa
    10081 atgtggaact gagtggatat gctgtttttg tctgttaaac agagaagctg gctgttatcc
    10141 actgagaagc gaacgaaaca gtcgggaaaa tctcccatta tcgtagagat ccgcattatt
    10201 aatctcagga gcctgtgtag cgtttatagg aagtagtgtt ctgtcatgat gcctgcaagc
    10261 ggtaacgaaa acgatttgaa tatgccttca ggaacaatag aaatcttcgt gcggtgttac
    10321 gttgaagtgg agcggattat gtcagcaatg gacagaacaa cctaatgaac acagaaccat
    10381 gatgtggtct gtccttttac agccagtagt gctcgccgca gtcgagcgac agggcgaagc
    10441 cctcggctgg ttgccctcgc cgctgggctg gcggccgtct atggccctgc aaacgcgcca
    10501 gaaacgccgt cgaagccgtg tgcgagacac cgcggccggc cgccggcgtt gtggatacct
    10561 cgcggaaaac ttggccctca ctgacagatg aggggcggac gttgacactt gaggggccga
    10621 ctcacccggc gcggcgttga cagatgaggg gcaggctcga tttcggccgg cgacgtggag
    10681 ctggccagcc tcgcaaatcg gcgaaaacgc ctgattttac gcgagtttcc cacagatgat
    10741 gtggacaagc ctggggataa gtgccctgcg gtattgacac ttgaggggcg cgactactga
    10801 cagatgaggg gcgcgatcct tgacacttga ggggcagagt gctgacagat gaggggcgca
    10861 cctattgaca tttgaggggc tgtccacagg cagaaaatcc agcatttgca agggtttccg
    10921 cccgtttttc ggccaccgct aacctgtctt ttaacctgct tttaaaccaa tatttataaa
    10981 ccttgttttt aaccagggct gcgccctgtg cgcgtgaccg cgcacgccga aggggggtgc
    11041 ccccccttct cgaaccctcc cggtcgagtg agcgaggaag caccagggaa cagcacttat
    11101 atattctgct tacacacgat gcctgaaaaa acttcccttg gggttatcca cttatccacg
    11161 gggatatttt tataattatt ttttttatag tttttagatc ttctttttta gagcgccttg
    11221 taggccttta tccatgctgg ttctagagaa ggtgttgtga caaattgccc tttcagtgtg
    11281 acaaatcacc ctcaaatgac agtcctgtct gtgacaaatt gcccttaacc ctgtgacaaa
    11341 ttgccctcag aagaagctgt tttttcacaa agttatccct gcttattgac tcttttttat
    11401 ttagtgtgac aatctaaaaa cttgtcacac ttcacatgga tctgtcatgg cggaaacagc
    11461 ggttatcaat cacaagaaac gtaaaaatag cccgcgaatc gtccagtcaa acgacctcac
    11521 tgaggcggca tatagtctct cccgggatca aaaacgtatg ctgtatctgt tcgttgacca
    11581 gatcagaaaa tctgatggca ccctacagga acatgacggt atctgcgaga tccatgttgc
    11641 taaatatgct gaaatattcg gattgacctc tgcggaagcc agtaaggata tacggcaggc
    11701 attgaagagt ttcgcgggga aggaagtggt tttttatcgc cctgaagagg atgccggcga
    11761 tgaaaaaggc tatgaatctt ttccttggtt tatcaaacgt gcgcacagtc catccagagg
    11821 gctttacagt gtacatatca acccatatct cattcccttc tttatcgggt tacagaaccg
    11881 gtttacgcag tttcggctta gtgaaacaaa agaaatcacc aatccgtatg ccatgcgttt
    11941 atacgaatcc ctgtgtcagt atcgtaagcc ggatggctca ggcatcgtct ctctgaaaat
    12001 cgactggatc atagagcgtt accagctgcc tcaaagttac cagcgtatgc ctgacttccg
    12061 ccgccgcttc ctgcaggtct gtgttaatga gatcaacagc agaactccaa tgcgcctctc
    12121 atacattgag aaaaagaaag gccgccagac gactcatatc gtattttcct tccgcgatat
    12181 cacttccatg acgacaggat agtctgaggg ttatctgtca cagatttgag ggtggttcgt
    12241 cacatttgtt ctgacctact gagggtaatt tgtcacagtt ttgctgtttc cttcagcctg
    12301 catggatttt ctcatacttt ttgaactgta atttttaagg aagccaaatt tgagggcagt
    12361 ttgtcacagt tgatttcctt ctctttccct tcgtcatgtg acttgatatc gggggttagt
    12421 tcgtcatcat tgatgagggt tgattatcac agtttattac tctgaattgg ctatccgcgt
    12481 gtgtacctct acctggagtt tttcccacgg tggatatttc ttcttgcgct gagcgtaaga
    12541 gctatctgac agaacagttc ttctttgctt cctcgccagt tcgctcgcta tgctcggtta
    12601 cacggctgcg gcgagcgcta gtgataataa gtgactgagg tatgtgctct tcttatctcc
    12661 ttttgtagtg ttgctcttat tttaaacaac tttgcggttt tttgatgact ttgcgatttt
    12721 gttgttgctt tgcagtaaat tgcaagattt aataaaaaaa cgcaaagcaa tgattaaagg
    12781 atgttcagaa tgaaactcat ggaaacactt aaccagtgca taaacgctgg tcatgaaatg
    12841 acgaaggcta tcgccattgc acagtttaat gatgacagcc cggaagcgag gaaaataacc
    12901 cggcgctgga gaataggtga agcagcggat ttagttgggg tttcttctca ggctatcaga
    12961 gatgccgaga aagcagggcg actaccgcac ccggatatgg aaattcgagg acgggttgag
    13021 caacgtgttg gttatacaat tgaacaaact aatcatatgc gtgatgtgtt tggtacgcga
    13081 ttgcgacgtg ctgaagacgt atttccaccg gtgatcgggg ttgctgccca taaaggtggc
    13141 gtttacaaaa cctcagtttc tgttcatctt gctcaggatc tggctctgaa ggggctacgt
    13201 gttttgctcg tggaaggtaa cgacccccag ggaacagcct caatgtatca cggatgggta
    13261 ccagatcttc atattcatgc agaagacact ctcctgcctt tctatcttgg ggaaaaggac
    13321 gatgtcactt atgcaataaa gcccacttgc tggccggggc ttgacattat tccttcctgt
    13381 ctggctctgc accgtattga aactgagtta atgggcaaat ttgatgaagg taaactgccc
    13441 accgatccac acctgatgct ccgactggcc attgaaactg ttgctcatga ctatgatgtc
    13501 atagttattg acagcgcgcc taacctgggt atcggcacga ttaatgtcgt atgtgctgct
    13561 gatgtgctga ttgttcccac gcctgctgag ttgtttgact acacctccgc actgcagttt
    13621 ttcgatatgc ttcgtgatct gctcaagaac gttgatctta aagggttcga gcctgatgta
    13681 cgtattttgc ttaccaaata cagcaatagt aatggctctc agtccccgtg gatggaggag
    13741 caaattcggg atgcctgggg aagcatggtt ctaaaaaatg ttgtacgtga aacggatgaa
    13801 gttggtaaag gtcagatccg gatgagaact gtttttgaac aggccattga tcaacgctct
    13861 tcaactggtg cctggagaaa tgctctttct atttgggaac ctgtctgcaa tgaaattttc
    13921 gatcgtctga ttaaaccacg ctgggagatt agataatgaa gcgtgcgcct gttattccaa
    13981 aacatacgct caatactcaa ccggttgaag atacttcgtt atcgacacca gctgccccga
    14041 tggtggattc gttaattgcg cgcgtaggag taatggctcg cggtaatgcc attactttgc
    14101 ctgtatgtgg tcgggatgtg aagtttactc ttgaagtgct ccggggtgat agtgttgaga
    14161 agacctctcg ggtatggtca ggtaatgaac gtgaccagga gctgcttact gaggacgcac
    14221 tggatgatct catcccttct tttctactga ctggtcaaca gacaccggcg ttcggtcgaa
    14281 gagtatctgg tgtcatagaa attgccgatg ggagtcgccg tcgtaaagct gctgcactta
    14341 ccgaaagtga ttatcgtgtt ctggttggcg agctggatga tgagcagatg gctgcattat
    14401 ccagattggg taacgattat cgcccaacaa gtgcttatga acgtggtcag cgttatgcaa
    14461 gccgattgca gaatgaattt gctggaaata tttctgcgct ggctgatgcg gaaaatattt
    14521 cacgtaagat tattacccgc tgtatcaaca ccgccaaatt gcctaaatca gttgttgctc
    14581 ttttttctca ccccggtgaa ctatctgccc ggtcaggtga tgcacttcaa aaagccttta
    14641 cagataaaga ggaattactt aagcagcagg catctaacct tcatgagcag aaaaaagctg
    14701 gggtgatatt tgaagctgaa gaagttatca ctcttttaac ttctgtgctt aaaacgtcat
    14761 ctgcatcaag aactagttta agctcacgac atcagtttgc tcctggagcg acagtattgt
    14821 ataagggcga taaaatggtg cttaacctgg acaggtctcg tgttccaact gagtgtatag
    14881 agaaaattga ggccattctt aaggaacttg aaaagccagc accctgatgc gaccacgttt
    14941 tagtctacgt ttatctgtct ttacttaatg tcctttgtta caggccagaa agcataactg
    15001 gcctgaatat tctctctggg cccactgttc cacttgtatc gtcggtctga taatcagact
    15061 gggaccatgg tcccactcgt atcgtcggtc tgattattag tctgggacca cggtcccact
    15121 cgtatcgtcg gtctgattat tagtctggga ccacggtccc actcgtatcg tcggtctgat
    15181 aatcagactg ggaccacggt cccactcgta tcgtcggtct gattattagt ctgggaccat
    15241 ggtcccactc gtatcgtcgg tctgattatt agtctgggac cacggtccca ctcgtatcgt
    15301 cggtctgatt attagtctgg aaccacggtc ccactcgtat cgtcggtctg attattagtc
    15361 tgggaccacg gtcccactcg tatcgtcggt ctgattatta gtctgggacc acgatcccac
    15421 tcgtgttgtc ggtctgatta tcggtctggg accacggtcc cacttgtatt gtcgatcaga
    15481 ctatcagcgt gagactacga ttccatcaat gcctgtcaag ggcaagtatt gacatgtcgt
    15541 cgtaacctgt agaacggagt aacctcggtg tgcggttgta tgcctgctgt ggattgctgc
    15601 tgtgtcctgc ttatccacaa cattttgcgc acggttatgt ggacaaaata cctggttacc
    15661 caggccgtgc cggcacgtta accgggctgc atccgatgca agtgtgtcgc tgtcgacgag
//
