LOCUS       sequence-417141-       11422 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Reporter gene expression in B. dendrobatidis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11422)
  AUTHORS   Kalinka E, Brody SM, Swafford AJM, Medina EM, Fritz-Laylin LK
  TITLE     Genetic transformation of the frog-killing chytrid fungus 
            Batrachochytrium dendrobatidis.
  JOURNAL   Proc Natl Acad Sci U S A. 2024 Jan 23;121(4):e2317928121. doi: 
            10.1073/pnas.2317928121. Epub 2024 Jan 18.
  PUBMED    38236738
REFERENCE   2  (bases 1 to 11422)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2024-01-23"; volume: "121(4):e2317928121. 
            doi"; pages: " 10.1073/pnas.2317928121. Epub 2024 Jan 18"
COMMENT     Sequence Label: pEK20
FEATURES             Location/Qualifiers
     source          1..11422
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..25
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     primer_bind     206..228
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     220..237
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     221..237
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             525..1550
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             525..1550
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             1566..2081
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     terminator      2103..2290
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2158..2182)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(2305..2321)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2305..2321)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2318..2340)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2329..2345
                     /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(2353..2359,2360..2377,2378..2383))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2398..2419
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    2596..2620
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     primer_bind     complement(2902..2921)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2913..3728
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             2913..3728
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2986..3005)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3823..4411
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4312..4331
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4565..4582
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4597..4737
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      5081..5275
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     CDS             complement(5341..6414)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     CDS             complement(6843..7472)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     rep_origin      8633..9975
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(9596..9643)
                     /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)."
     primer_bind     10016..10034
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        10237..10452
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             10453..11256
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
ORIGIN
        1 gtttacccgc caatatatcc tgtcaaacac tgatagttta aactgaaggc gggaaacgac
       61 aatctgatcc aagctcaagc tcattcgcca ttcaggctgc gcaactgttg ggaagggcga
      121 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
      181 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc
      241 aagcttagta gtgtataaat gaagatgatt acaaagtcga gtttattgac aaggaattta
      301 agaatgataa gaattagaac accaagaata acattcaaga ttagatctaa ttctagagtc
      361 taaatgcatt ctggctaatc catcaacaac agaaacaagc atcaataatg tggctgaagc
      421 attcagggtt gtgtgcgggc agcagcaatc aactgtcaat ccaacaaatt ccattctttc
      481 caactcgctc acaattctat tattacccaa catcgcatta attaatgggt aaaaagcctg
      541 aactcaccgc gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc
      601 tgatgcagct ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg
      661 gatatgtcct gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc
      721 ggcactttgc atcggccgcg ctcccgattc cggaagtgct tgacattggg gaattcagcg
      781 agagcctgac ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg
      841 aaaccgaact gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg
      901 ccgatcttag ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca
      961 ctacatggcg tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg
     1021 tgatggacga caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg
     1081 ccgaggactg ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc
     1141 tgacggacaa tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt
     1201 cccaatacga ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc
     1261 agacgcgcta cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt
     1321 atatgctccg cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg
     1381 atgcagcttg ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg
     1441 ggcgtacaca aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt gtagaagtac
     1501 tcgccgatag tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa ggaggaggag
     1561 gatccatggt cttcacactc gaagatttcg ttggggactg gcgacagaca gccggctaca
     1621 acctggacca agtccttgaa cagggaggtg tgtccagttt gtttcagaat ctcggggtgt
     1681 ccgtaactcc gatccaaagg attgtcctga gcggtgaaaa tgggctgaag atcgacatcc
     1741 atgtcatcat cccgtatgaa ggtctgagcg gcgaccaaat gggccagatc gaaaaaattt
     1801 ttaaggtggt gtaccctgtg gatgatcatc actttaaggt gatcctgcac tatggcacac
     1861 tggtaatcga cggggttacg ccgaacatga tcgactattt cggacggccg tatgaaggca
     1921 tcgccgtgtt cgacggcaaa aagatcactg taacagggac cctgtggaac ggcaacaaaa
     1981 ttatcgacga gcgcctgatc aaccccgacg gctccctgct gttccgagta accatcaacg
     2041 gagtgaccgg ctggcggctg tgcgaacgca ttctggcgta aaggcgcgcc acttctaaat
     2101 aagcgaattt cttatgattt atgattttta ttattaaata agttataaaa aaaataagtg
     2161 tatacaaatt ttaaagtgac tcttaggttt taaaacgaaa attcttattc ttgagtaact
     2221 ctttcctgta ggtcaggttg ctttctcagg tatagtatga ggtcgctctt attgaccaca
     2281 cctctaccgg gaattcgtaa tcatgtcata gctgtttcct gtgtgaaatt gttatccgct
     2341 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     2401 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     2461 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2521 ctagagcaat tcggcgttaa ttcagtacat taaaaacgtc cgcaatgtgt tattaagttg
     2581 tctaagcgtc aatttgttta caccacaata tatcctgcca ccagccagcc aacagctccc
     2641 cgaccggcag ctcggcacaa aatcaccact cgatacaggc agcccatcag tccgggacgg
     2701 cgtcagcggg agagccgttg taaggcggca gactttgctc atgttaccga tgctattcgg
     2761 aagaacggca actaagctgc cgggtttgaa acacggatga tctcgcggag ggtagcatgt
     2821 tgattgtaac gatgacagag cgttgctgcc tgtgatccag atcatgaaca ataaaactgt
     2881 ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtctt
     2941 gctctaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc
     3001 gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc
     3061 cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg
     3121 tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta
     3181 ctcctgatga tgcatggtta ctcaccactg cgatccccgg gaaaacagca ttccaggtat
     3241 tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc
     3301 ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg
     3361 ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc
     3421 gtaatggctg gcctgttgaa caagtctgga aagaaatgca taaacttttg ccattctcac
     3481 cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga
     3541 aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg
     3601 ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa
     3661 aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt
     3721 ttttctaaga attaattcat gatcctgcat gaccaaaatc ccttaacgtg agttttcgtt
     3781 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     3841 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     3901 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     3961 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4021 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4081 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4141 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4201 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4261 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4321 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4381 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4441 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4501 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     4561 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     4621 gcatctgtgc ggtatttcac accgcatatg gtgcactctc agtacaatct gctctgatgc
     4681 cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc
     4741 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     4801 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     4861 ccgaaacgcg cgaggcaggg tgccttgatg tgggcgccgg cggtcgagtg gcgacggcgc
     4921 ggcttgtccg cgccctggta gattgcctgg ccgtaggcca gccatttttg agcggccagc
     4981 ggccgcgata ggccgacgcg aagcggcggg gcgtagggag cgcagcgacc gaagggtagg
     5041 cgctttttgc agctcttcgg ctgtgcgctg gccagacagt tatgcacagg ccaggcgggt
     5101 tttaagagtt ttaataagtt ttaaagagtt ttaggcggaa aaatcgcctt ttttctcttt
     5161 tatatcagtc acttacatgt gtgaccggtt cccaatgtac ggctttgggt tcccaatgta
     5221 cgggttccgg ttcccaatgt acggctttgg gttcccaatg tacgtgctat ccacaggaaa
     5281 gagacctttt cgaccttttt cccctgctag ggcaatttgc cctagcatct gctccgtaca
     5341 ttaggaaccg gcggatgctt cgccctcgat caggttgcgg tagcgcatga ctaggatcgg
     5401 gccagcctgc cccgcctcct ccttcaaatc gtactccggc aggtcatttg acccgatcag
     5461 cttgcgcacg gtgaaacaga acttcttgaa ctctccggcg ctgccactgc gttcgtagat
     5521 cgtcttgaac aaccatctgg cttctgcctt gcctgcggcg cggcgtgcca ggcggtagag
     5581 aaaacggccg atgccgggat cgatcaaaaa gtaatcgggg tgaaccgtca gcacgtccgg
     5641 gttcttgcct tctgtgatct cgcggtacat ccaatcagct agctcgatct cgatgtactc
     5701 cggccgcccg gtttcgctct ttacgatctt gtagcggcta atcaaggctt caccctcgga
     5761 taccgtcacc aggcggccgt tcttggcctt cttcgtacgc tgcatggcaa cgtgcgtggt
     5821 gtttaaccga atgcaggttt ctaccaggtc gtctttctgc tttccgccat cggctcgccg
     5881 gcagaacttg agtacgtccg caacgtgtgg acggaacacg cggccgggct tgtctccctt
     5941 cccttcccgg tatcggttca tggattcggt tagatgggaa accgccatca gtaccaggtc
     6001 gtaatcccac acactggcca tgccggccgg ccctgcggaa acctctacgt gcccgtctgg
     6061 aagctcgtag cggatcacct cgccagctcg tcggtcacgc ttcgacagac ggaaaacggc
     6121 cacgtccatg atgctgcgac tatcgcgggt gcccacgtca tagagcatcg gaacgaaaaa
     6181 atctggttgc tcgtcgccct tgggcggctt cctaatcgac ggcgcaccgg ctgccggcgg
     6241 ttgccgggat tctttgcgga ttcgatcagc ggccgcttgc cacgattcac cggggcgtgc
     6301 ttctgcctcg atgcgttgcc gctgggcggc ctgcgcggcc ttcaacttct ccaccaggtc
     6361 atcacccagc gccgcgccga tttgtaccgg gccggatggt ttgcgaccgc tcacgccgat
     6421 tcctcgggct tgggggttcc agtgccattg cagggccggc agacaaccca gccgcttacg
     6481 cctggccaac cgcccgttcc tccacacatg gggcattcca cggcgtcggt gcctggttgt
     6541 tcttgatttt ccatgccgcc tcctttagcc gctaaaattc atctactcat ttattcattt
     6601 gctcatttac tctggtagct gcgcgatgta ttcagatagc agctcggtaa tggtcttgcc
     6661 ttggcgtacc gcgtacatct tcagcttggt gtgatcctcc gccggcaact gaaagttgac
     6721 ccgcttcatg gctggcgtgt ctgccaggct ggccaacgtt gcagccttgc tgctgcgtgc
     6781 gctcggacgg ccggcactta gcgtgtttgt gcttttgctc attttctctt tacctcatta
     6841 actcaaatga gttttgattt aatttcagcg gccagcgcct ggacctcgcg ggcagcgtcg
     6901 ccctcgggtt ctgattcaag aacggttgtg ccggcggcgg cagtgcctgg gtagctcacg
     6961 cgctgcgtga tacgggactc aagaatgggc agctcgtacc cggccagcgc ctcggcaacc
     7021 tcaccgccga tgcgcgtgcc tttgatcgcc cgcgacacga caaaggccgc ttgtagcctt
     7081 ccatccgtga cctcaatgcg ctgcttaacc agctccacca ggtcggcggt ggcccatatg
     7141 tcgtaagggc ttggctgcac cggaatcagc acgaagtcgg ctgccttgat cgcggacaca
     7201 gccaagtccg ccgcctgggg cgctccgtcg atcactacga agtcgcgccg gccgatggcc
     7261 ttcacgtcgc ggtcaatcgt cgggcggtcg atgccgacaa cggttagcgg ttgatcttcc
     7321 cgcacggccg cccaatcgcg ggcactgccc tggggatcgg aatcgactaa cagaacatcg
     7381 gccccggcga gttgcagggc gcgggctaga tgggttgcga tggtcgtctt gcctgacccg
     7441 cctttctggt taagtacagc gataaccttc atgcgttccc cttgcgtatt tgtttattta
     7501 ctcatcgcat catatacgca gcgaccgcat gacgcaagct gttttactca aatacacatc
     7561 acctttttag acggcggcgc tcggtttctt cagcggccaa gctggccggc caggccgcca
     7621 gcttggcatc agacaaaccg gccaggattt catgcagccg cacggttgag acgtgcgcgg
     7681 gcggctcgaa cacgtacccg gccgcgatca tctccgcctc gatctcttcg gtaatgaaaa
     7741 acggttcgtc ctggccgtcc tggtgcggtt tcatgcttgt tcctcttggc gttcattctc
     7801 ggcggccgcc agggcgtcgg cctcggtcaa tgcgtcctca cggaaggcac cgcgccgcct
     7861 ggcctcggtg ggcgtcactt cctcgctgcg ctcaagtgcg cggtacaggg tcgagcgatg
     7921 cacgccaagc agtgcagccg cctctttcac ggtgcggcct tcctggtcga tcagctcgcg
     7981 ggcgtgcgcg atctgtgccg gggtgagggt agggcggggg ccaaacttca cgcctcgggc
     8041 cttggcggcc tcgcgcccgc tccgggtgcg gtcgatgatt agggaacgct cgaactcggc
     8101 aatgccggcg aacacggtca acaccatgcg gccggccggc gtggtggtgt cggcccacgg
     8161 ctctgccagg ctacgcaggc ccgcgccggc ctcctggatg cgctcggcaa tgtccagtag
     8221 gtcgcgggtg ctgcgggcca ggcggtctag cctggtcact gtcacaacgt cgccagggcg
     8281 taggtggtca agcatcctgg ccagctccgg gcggtcgcgc ctggtgccgg tgatcttctc
     8341 ggaaaacagc ttggtgcagc cggccgcgtg cagttcggcc cgttggttgg tcaagtcctg
     8401 gtcgtcggtg ctgacgcggg catagcccag caggccagcg gcggcgctct tgttcatggc
     8461 gtaatgtctc cggttctagt cgcaagtatt ctactttatg cgactaaaac acgcgacaag
     8521 aaaacgccag gaaaagggca gggcggcagc ctgtcgcgta acttaggact tgtgcgacat
     8581 gtcgttttca gaagacggct gcactgaacg tcagaaggaa aagtgccacc tgaacgaagc
     8641 atctgtgctt cattttgtag aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa
     8701 gaatctgagc tgcattttta cagaacagaa atgcaacgcg aaagcgctat tttaccaacg
     8761 aagaatctgt gcttcatttt tgtaaaacaa aaatgcaacg cgagagcgct aatttttcaa
     8821 acaaagaatc tgagctgcat ttttacagaa cagaaatgca acgcgagagc gctattttac
     8881 caacaaagaa tctatacttc ttttttgttc tacaaaaatg catcccgaga gcgctatttt
     8941 tctaacaaag catcttagat tacttttttt ctcctttgtg cgctctataa tgcagtctct
     9001 tgataacttt ttgcactgta ggtccgttaa ggttagaaga aggctacttt ggtgtctatt
     9061 ttctcttcca taaaaaaagc ctgactccac ttcccgcgtt tactgattac tagcgaagct
     9121 gcgggtgcat tttttcaaga taaaggcatc cccgattata ttctataccg atgtggattg
     9181 cgcatacttt gtgaacagaa agtgatagcg ttgatgattc ttcattggtc agaaaattat
     9241 gaacggtttc ttctattttg tctctatata ctacgtatag gaaatgttta cattttcgta
     9301 ttgttttcga ttcactctat gaatagttct tactacaatt tttttgtcta aagagtaata
     9361 ctagagataa acataaaaaa tgtagaggtc gagtttagat gcaagttcaa ggagcgaaag
     9421 gtggatgggt aggttatata gggatatagc acagagatat atagcaaaga gatacttttg
     9481 agcaatgttt gtggaagcgg tattcgcaat attttagtag ctcgttacag tccggtgcgt
     9541 ttttggtttt ttgaaagtgc gtcttcagag cgcttttggt tttcaaaagc gctctgaagt
     9601 tcctatactt tctagagaat aggaacttcg gaataggaac ttcaaagcgt ttccgaaaac
     9661 gagcgcttcc gaaaatgcaa cgcgagctgc gcacatacag ctcactgttc acgtcgcacc
     9721 tatatctgcg tgttgcctgt atatatatat acatgagaag aacggcatag tgcgtgttta
     9781 tgcttaaatg cgtacttata tgcgtctatt tatgtaggat gaaaggtagt ctagtacctc
     9841 ctgtgatatt atcccattcc atgcggggta tcgtatgctt ccttcagcac taccctttag
     9901 ctgttctata tgctgccact cctcaattgg attagtctca tccttcaatg ctatcatttc
     9961 ctttgatatt ggatcatact aagaaaccat tattatcatg acattaacct ataaaaatag
    10021 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
    10081 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
    10141 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
    10201 agagcagatt gtactgagag tgcaccatac caccttttca attcatcatt ttttttttat
    10261 tctttttttt gatttcggtt tccttgaaat ttttttgatt cggtaatctc cgaacagaag
    10321 gaagaacgaa ggaaggagca cagacttaga ttggtatata tacgcatatg tagtgttgaa
    10381 gaaacatgaa attgcccagt attcttaacc caactgcaca gaacaaaaac ctgcaggaaa
    10441 cgaagataaa tcatgtcgaa agctacatat aaggaacgtg ctgctactca tcctagtcct
    10501 gttgctgcca agctatttaa tatcatgcac gaaaagcaaa caaacttgtg tgcttcattg
    10561 gatgttcgta ccaccaagga attactggag ttagttgaag cattaggtcc caaaatttgt
    10621 ttactaaaaa cacatgtgga tatcttgact gatttttcca tggagggcac agttaagccg
    10681 ctaaaggcat tatccgccaa gtacaatttt ttactcttcg aagacagaaa atttgctgac
    10741 attggtaata cagtcaaatt gcagtactct gcgggtgtat acagaatagc agaatgggca
    10801 gacattacga atgcacacgg tgtggtgggc ccaggtattg ttagcggttt gaagcaggcg
    10861 gcagaagaag taacaaagga acctagaggc cttttgatgc tagcagaatt gtcatgcaag
    10921 ggctccctat ctactggaga atatactaag ggtactgttg acattgcgaa gagcgacaaa
    10981 gattttgtta tcggctttat tgctcaaaga gacatgggtg gaagagatga aggttacgat
    11041 tggttgatta tgacacccgg tgtgggttta gatgacaagg gagacgcatt gggtcaacag
    11101 tatagaaccg tggatgatgt ggtctctaca ggatctgaca ttattattgt tggaagagga
    11161 ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa agcaggctgg
    11221 gaagcatatt tgagaagatg cggccagcaa aactaaaaaa ctgtattata agtaaatgca
    11281 tgtatactaa actcacaaat tagagcttca atttaattat atcagttatt acctggcatg
    11341 cacatacaaa tggacgaacg gataaacctt ttcacgccct tttaaatatc cgattattct
    11401 aataaacgct cttttctctt ag
//
