LOCUS       sequence-386424-       10918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express APOVEC1-nCas9(D10A)-UGI using aTC inducible system in 
            Methanotroph or E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10918)
  AUTHORS   Jeong J, Kim TH, Jang N, Ko M, Kim SK, Baek JI, Emelianov G, Rha E, 
            Kwon KK, Kim H, Lee EY, Lee DH, Lee H, Lee SG
  TITLE     A highly efficient and versatile genetic engineering toolkit for a 
            methanotroph-based biorefinery
  JOURNAL   Chem Eng J, 2023
REFERENCE   2  (bases 1 to 10918)
  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: "Chem Eng J,
            2023"
COMMENT     Sequence Label: pAWPSG-Ptet-nCas9-BE
FEATURES             Location/Qualifiers
     source          1..10918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      22..653
                     /label=oriV
                     /note="incP origin of replication"
     terminator      complement(917..1011)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1042..1668)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     1573..1592
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        1684..1739
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    1720..1738
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             1755..1777
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1785..2471
                     /codon_start=1
                     /product="cytidine deaminase (C to U editing enzyme) from 
                     rat"
                     /label=APOBEC-1
                     /note="can use ssDNA as a substrate (Komor et al., 2016)"
                     /translation="MSSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWG
                     GRHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFL
                     SRYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSN
                     EAHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHI
                     LWATGLK"
     CDS             join(2520..2543,2544..2546,2547..6620)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             6633..6884
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     terminator      6908..6979
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6995..7022
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             7090..7119
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             7135..7152
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7208..7225)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      7378..7424
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      7496..7582
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(7735..8550)
                     /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             complement(7735..8550)
                     /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     8458..8477
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     8542..8561
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(8854..10002)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     CDS             complement(10274..10645)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSL*AYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            10678..10787
                     /note="incP origin of transfer"
ORIGIN
        1 aaagatccgt cgaccctttc cgacgctcac cgggctggtt gccctcgccg ctgggctggc
       61 ggccgtctat ggccctgcaa acgcgccaga aacgccgtcg aagccgtgtg cgagacaccg
      121 cggccgccgg cgttgtggat acctcgcgga aaacttggcc ctcactgaca gatgaggggc
      181 ggacgttgac acttgagggg ccgactcacc cggcgcggcg ttgacagatg aggggcaggc
      241 tcgatttcgg ccggcgacgt ggagctggcc agcctcgcaa atcggcgaaa acgcctgatt
      301 ttacgcgagt ttcccacaga tgatgtggac aagcctgggg ataagtgccc tgcggtattg
      361 acacttgagg ggcgcgacta ctgacagatg aggggcgcga tccttgacac ttgaggggca
      421 gagtgctgac agatgagggg cgcacctatt gacatttgag gggctgtcca caggcagaaa
      481 atccagcatt tgcaagggtt tccgcccgtt tttcggccac cgctaacctg tcttttaacc
      541 tgcttttaaa ccaatattta taaaccttgt ttttaaccag ggctgcgccc tgtgcgcgtg
      601 accgcgcacg ccgaaggggg gtgccccccc ttctcgaacc ctcccggccc gctaacgcgg
      661 gcctcccatc cccccagggg ctgcgcccct cggccgcgaa cggcctcacc ccaaaaatgg
      721 cagccaagct gacccgctag ggacgtgaag tcgattcctt cgatggttag caatcaaaga
      781 ctcagagtgc tgtctggagc gtgaatctaa cggtacgtat ctcgattgct cggtcgctat
      841 tcgcactctg cgaaagttcg taccgctcat tcactaggtt gcgaaaattt aaatcgtaat
      901 tattggggac ccctggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga
      961 acaaatccag atggagttct gaggtcatta ctggatctat caacaggagt ccaagactag
     1021 tcgccagggt tttcccagtc attaagaccc actttcacat ttaagttgtt tttctaatcc
     1081 gcatatgatc aattcaaggc cgaataagaa ggctggctct gcaccttggt gatcaaataa
     1141 ttcgatagct tgtcgtaata atggcggcat actatcagta gtaggtgttt ccctttcttc
     1201 tttagcgact tgatgctctt gatcttccaa tacgcaacct aaagtaaaat gccccacagc
     1261 gctgagtgca tataatgcat tctctagtga aaaaccttgt tggcataaaa aggctaattg
     1321 attttcgaga gtttcatact gtttttctgt aggccgtgta cctaaatgta cttttgctcc
     1381 atcgcgatga cttagtaaag cacatctaaa acttttagcg ttattacgta aaaaatcttg
     1441 ccagctttcc ccttctaaag ggcaaaagtg agtatggtgc ctatctaaca tctcaatggc
     1501 taaggcgtcg agcaaagccc gcttattttt tacatgccaa tacaatgtag gctgctctac
     1561 acctagcttc tgggcgagtt tacgggttgt taaaccttcg attccgacct cattaagcag
     1621 ctctaatgcg ctgttaatca ctttactttt atctaatcta gacatcatta attcctaatt
     1681 tttgttgaca ctctatcgtt gatagagtta ttttaccact ccctatcagt gatagagaaa
     1741 agtctagaaa taattttgtt taactttaag aaggagatat acatatgagc tcagagactg
     1801 gcccagtggc tgtggacccc acattgagac ggcggatcga gccccatgag tttgaggtat
     1861 tcttcgatcc gagagagctc cgcaaggaga cctgcctgct ttacgaaatt aattgggggg
     1921 gccggcactc catttggcga catacatcac agaacactaa caagcacgtc gaagtcaact
     1981 tcatcgagaa gttcacgaca gaaagatatt tctgtccgaa cacaaggtgc agcattacct
     2041 ggtttctcag ctggagccca tgcggcgaat gtagtagggc catcactgaa ttcctgtcaa
     2101 ggtatcccca cgtcactctg tttatttaca tcgcaaggct gtaccaccac gctgaccccc
     2161 gcaatcgaca aggcctgcgg gatttgatct cttcaggtgt gactatccaa attatgactg
     2221 agcaggagtc aggatactgc tggagaaact ttgtgaatta tagcccgagt aatgaagccc
     2281 actggcctag gtatccccat ctgtgggtac gactgtacgt tcttgaactg tactgcatca
     2341 tactgggcct gcctccttgt ctcaacattc tgagaaggaa gcagccacag ctgacattct
     2401 ttaccatcgc tcttcagtct tgtcattacc agcgactgcc cccacacatt ctctgggcca
     2461 ccgggttgaa aagcggcagc gagactcccg ggacctcaga gtccgccaca cccgaaagtg
     2521 ataagaaata ctcaataggc ttagctatcg gcacaaatag cgtcggatgg gcggtgatca
     2581 ctgatgaata taaggttccg tctaaaaagt tcaaggttct gggaaataca gaccgccaca
     2641 gtatcaaaaa aaatcttata ggggctcttt tatttgacag tggagagaca gcggaagcga
     2701 ctcgtctcaa acggacagct cgtagaaggt atacacgtcg gaagaatcgt atttgttatc
     2761 tacaggagat tttttcaaat gagatggcga aagtagatga tagtttcttt catcgacttg
     2821 aagagtcttt tttggtggaa gaagacaaga agcatgaacg tcatcctatt tttggaaata
     2881 tagtagatga agttgcttat catgagaaat atccaactat ctatcatctg cgaaaaaaat
     2941 tggtagattc tactgataaa gcggatttgc gcttaatcta tttggcctta gcgcatatga
     3001 ttaagtttcg tggtcatttt ttgattgagg gagatttaaa tcctgataat agtgatgtgg
     3061 acaaactatt tatccagttg gtacaaacct acaatcaatt atttgaagaa aaccctatta
     3121 acgcaagtgg agtagatgct aaagcgattc tttctgcacg attgagtaaa tcaagacgat
     3181 tagaaaatct cattgctcag ctccccggtg agaagaaaaa tggcttattt gggaatctca
     3241 ttgctttgtc attgggtttg acccctaatt ttaaatcaaa ttttgatttg gcagaagatg
     3301 ctaaattaca gctttcaaaa gatacttacg atgatgattt agataattta ttggcgcaaa
     3361 ttggagatca atatgctgat ttgtttttgg cagctaagaa tttatcagat gctattttac
     3421 tttcagatat cctaagagta aatactgaaa taactaaggc tcccctatca gcttcaatga
     3481 ttaaacgcta cgatgaacat catcaagact tgactctttt aaaagcttta gttcgacaac
     3541 aacttccaga aaagtataaa gaaatctttt ttgatcaatc aaaaaacgga tatgcaggtt
     3601 atattgatgg gggagctagc caagaagaat tttataaatt tatcaaacca attttagaaa
     3661 aaatggatgg tactgaggaa ttattggtga aactaaatcg tgaagatttg ctgcgcaagc
     3721 aacggacctt tgacaacggc tctattcccc atcaaattca cttgggtgag ctgcatgcta
     3781 ttttgagaag acaagaagac ttttatccat ttttaaaaga caatcgtgag aagattgaaa
     3841 aaatcttgac ttttcgaatt ccttattatg ttggtccatt ggcgcgtggc aatagtcgtt
     3901 ttgcatggat gactcggaag tctgaagaaa caattacccc atggaatttt gaagaagttg
     3961 tcgataaagg tgcttcagct caatcattta ttgaacgcat gacaaacttt gataaaaatc
     4021 ttccaaatga aaaagtacta ccaaaacata gtttgcttta tgagtatttt acggtttata
     4081 acgaattgac aaaggtcaaa tatgttactg aaggaatgcg aaaaccagca tttctttcag
     4141 gtgaacagaa gaaagccatt gttgatttac tcttcaaaac aaatcgaaaa gtaaccgtta
     4201 agcaattaaa agaagattat ttcaaaaaaa tagaatgttt tgatagtgtt gaaatttcag
     4261 gagttgaaga tagatttaat gcttcattag gtacctacca tgatttgcta aaaattatta
     4321 aagataaaga ttttttggat aatgaagaaa atgaagatat cttagaggat attgttttaa
     4381 cattgacctt atttgaagat agggagatga ttgaggaaag acttaaaaca tatgctcacc
     4441 tctttgatga taaggtgatg aaacagctta aacgtcgccg ttatactggt tggggacgtt
     4501 tgtctcgaaa attgattaat ggtattaggg ataagcaatc tggcaaaaca atattagatt
     4561 ttttgaaatc agatggtttt gccaatcgca attttatgca gctgatccat gatgatagtt
     4621 tgacatttaa agaagacatt caaaaagcac aagtgtctgg acaaggcgat agtttacatg
     4681 aacatattgc aaatttagct ggtagccctg ctattaaaaa aggtatttta cagactgtaa
     4741 aagttgttga tgaattggtc aaagtaatgg ggcggcataa gccagaaaat atcgttattg
     4801 aaatggcacg tgaaaatcag acaactcaaa agggccagaa aaattcgcga gagcgtatga
     4861 aacgaatcga agaaggtatc aaagaattag gaagtcagat tcttaaagag catcctgttg
     4921 aaaatactca attgcaaaat gaaaagctct atctctatta tctccaaaat ggaagagaca
     4981 tgtatgtgga ccaagaatta gatattaatc gtttaagtga ttatgatgtc gatcacattg
     5041 ttccacaaag tttccttaaa gacgattcaa tagacaataa ggtcttaacg cgttctgata
     5101 aaaatcgtgg taaatcggat aacgttccaa gtgaagaagt agtcaaaaag atgaaaaact
     5161 attggagaca acttctaaac gccaagttaa tcactcaacg taagtttgat aatttaacga
     5221 aagctgaacg tggaggtttg agtgaacttg ataaagctgg ttttatcaaa cgccaattgg
     5281 ttgaaactcg ccaaatcact aagcatgtgg cacaaatttt ggatagtcgc atgaatacta
     5341 aatacgatga aaatgataaa cttattcgag aggttaaagt gattacctta aaatctaaat
     5401 tagtttctga cttccgaaaa gatttccaat tctataaagt acgtgagatt aacaattacc
     5461 atcatgccca tgatgcgtat ctaaatgccg tcgttggaac tgctttgatt aagaaatatc
     5521 caaaacttga atcggagttt gtctatggtg attataaagt ttatgatgtt cgtaaaatga
     5581 ttgctaagtc tgagcaagaa ataggcaaag caaccgcaaa atatttcttt tactctaata
     5641 tcatgaactt cttcaaaaca gaaattacac ttgcaaatgg agagattcgc aaacgccctc
     5701 taatcgaaac taatggggaa actggagaaa ttgtctggga taaagggcga gattttgcca
     5761 cagtgcgcaa agtattgtcc atgccccaag tcaatattgt caagaaaaca gaagtacaga
     5821 caggcggatt ctccaaggag tcaattttac caaaaagaaa ttcggacaag cttattgctc
     5881 gtaaaaaaga ctgggatcca aaaaaatatg gtggttttga tagtccaacg gtagcttatt
     5941 cagtcctagt ggttgctaag gtggaaaaag ggaaatcgaa gaagttaaaa tccgttaaag
     6001 agttactagg gatcacaatt atggaaagaa gttcctttga aaaaaatccg attgactttt
     6061 tagaagctaa aggatataag gaagttaaaa aagacttaat cattaaacta cctaaatata
     6121 gtctttttga gttagaaaac ggtcgtaaac ggatgctggc tagtgccgga gaattacaaa
     6181 aaggaaatga gctggctctg ccaagcaaat atgtgaattt tttatattta gctagtcatt
     6241 atgaaaagtt gaagggtagt ccagaagata acgaacaaaa acaattgttt gtggagcagc
     6301 ataagcatta tttagatgag attattgagc aaatcagtga attttctaag cgtgttattt
     6361 tagcagatgc caatttagat aaagttctta gtgcatataa caaacataga gacaaaccaa
     6421 tacgtgaaca agcagaaaat attattcatt tatttacgtt gacgaatctt ggagctcccg
     6481 ctgcttttaa atattttgat acaacaattg atcgtaaacg atatacgtct acaaaagaag
     6541 ttttagatgc cactcttatc catcaatcca tcactggtct ttatgaaaca cgcattgatt
     6601 tgagtcagct aggaggtgac tctggtggtt ctactaatct gtcagatatt attgaaaagg
     6661 agaccggtaa gcaactggtt atccaggaat ccatcctcat gctcccagag gaggtggaag
     6721 aagtcattgg gaacaagccg gaaagcgata tactcgtgca caccgcctac gacgagagca
     6781 ccgacgagaa tgtcatgctt ctgactagcg acgcccctga atacaagcct tgggctctgg
     6841 tcatacagga tagcaacggt gagaacaaga ttaagatgct ctaactcgag taaggatctc
     6901 caggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg
     6961 tttgtcggtg aacgctctct actagagtca cactggctca ccttcgggtg ggcctttctg
     7021 cgtttatacc tagggatata ttccgcttcc tcggcgaccg gggcggccgc ctcgaggaag
     7081 cttgggcccg aacaaaaact catctcagaa gaggatctga atagcgccgt cgaccatcat
     7141 catcatcatc attgagttta aacggtctcc agcttggctg ttttggcgga tgagagaaga
     7201 ttttcagcct gatacagatt aaatcagaac gcagaagcgg tctgataaaa cagaatttgc
     7261 ctggcggcag tagcgcggtg gtcccacctg accccatgcc gaactcagaa gtgaaacgcc
     7321 gtagcgccga tggtagtgtg gggtctcccc atgcgagagt agggaactgc caggcatcaa
     7381 ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg
     7441 aactggcgcc caatacgcaa accgtgaaat tgttatccgc tttaattaaa ggcatcaaat
     7501 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     7561 cgctctcctg agtaggacaa atccgccgcc ctagacagct gggcgcgcca ttgtcgggaa
     7621 gatgcgtgat ctgatccttc cactcagcaa aagttcgatt tattcaacaa agccgccgtc
     7681 ccgtcaagtc agcgtaatgc tctgccagtg ttacaaccaa ttaaccaatt ctgattagaa
     7741 aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata
     7801 tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat
     7861 ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa
     7921 tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc
     7981 cggtgagaat ggcaaaagct tatgcatttc tttccagact tgttcaacag gccagccatt
     8041 acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg
     8101 agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa
     8161 ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc
     8221 taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg
     8281 agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct
     8341 gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc
     8401 tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc
     8461 gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctcga
     8521 gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc
     8581 agacagtttt attgttcatg atgatatatt tttatcttgt gcaatgtaac atcagagatt
     8641 ttgagacaca acgtggcttt cgacttccgg caagctatac gcgccctaga attgtcaatt
     8701 ttaatcctct gtttatcggc agttcgtaga gcgcgccgtg cgtcccgagc gatactgagc
     8761 gaagcaagtg cgtcgagcag tgcccgcttg ttcctgaaat gccagtaaag cgctggctgc
     8821 tgaaccccca gccggaactg accccacaag gccctagcgt ttgcaatgca ccaggtcatc
     8881 attgacccag gcgtgttcca ccaggccgct gcctcgcaac tcttcgcagg cttcgccgac
     8941 ctgctcgcgc cacttcttca cgcgggtgga atccgatccg cacatgaggc ggaaggtttc
     9001 cagcttgagc gggtacggct cccggtgcga gctgaaatag tcgaacatcc gtcgggccgt
     9061 cggcgacagc ttgcggtact tctcccatat gaatttcgtg tagtggtcgc cagcaaacag
     9121 cacgacgatt tcctcgtcga tcaggacctg gcaacgggac gttttcttgc cacggtccag
     9181 gacgcggaag cggtgcagca gcgacaccga ttccaggtgc ccaacgcggt cggacgtgaa
     9241 gcccatcgcc gtcgcctgta ggcgcgacag gcattcctcg gccttcgtgt aataccggcc
     9301 attgatcgac cagcccaggt cctggcaaag ctcgtagaac gtgaaggtga tcggctcgcc
     9361 gataggggtg cgcttcgcgt actccaacac ctgctgccac accagttcgt catcgtcggc
     9421 ccgcagctcg acgccggtgt aggtgatctt cacgtccttg ttgacgtgga aaatgacctt
     9481 gttttgcagc gcctcgcgcg ggattttctt gttgcgcgtg gtgaacaggg cagagcgggc
     9541 cgtgtcgttt ggcatcgctc gcatcgtgtc cggccacggc gcaatatcga acaaggaaag
     9601 ctgcatttcc ttgatctgct gcttcgtgtg tttcagcaac gcggcctgct tggcctcgct
     9661 gacctgtttt gccaggtcct cgccggcggt ttttcgcttc ttggtcgtca tagttcctcg
     9721 cgtgtcgatg gtcatcgact tcgccaaacc tgccgcctcc tgttcgagac gacgcgaacg
     9781 ctccacggcg gccgatggcg cgggcagggc agggggagcc agttgcacgc tgtcgcgctc
     9841 gatcttggcc gtagcttgct ggaccatcga gccgacggac tggaaggttt cgcggggcgc
     9901 acgcatgacg gtgcggcttg cgatggtttc ggcatcctcg gcggaaaacc ccgcgtcgat
     9961 cagttcttgc ctgtatgcct tccggtcaaa cgtccgattc attcaccctc cttgcgggat
    10021 tgccccgact cacgccgggg caatgtgccc ttattcctga tttgacccgc ctggtgcctt
    10081 ggtgtccaga taatccacct tatcggcaat gaagtcggtc ccgtagaccg tctggccgtc
    10141 cttctcgtac ttggtattcc gaatcttgcc ctgcacgaat accagctccg cgaagtcgct
    10201 cttcttgatg gagcgcatgg ggacgtgctt ggcaatcacg cgcacccccc ggccgtttta
    10261 gcggctaaaa aagtcatggc tctgccctcg ggcggaccac gcccatcatg accttgccaa
    10321 gctcgtcctg cttctcttcg atcttcgcca gcagggcgag gatcgtggca tcaccgaacc
    10381 gcgccgtgcg cgggtcgtcg gtgagccaga gtttcagcag gccgcccagg cggcccaggt
    10441 cgccattgat gcgggccagc tcgcggacgt gctcatagtc cacgacgccc gtgattttgt
    10501 agccctggcc gacggccagc aggtaggcct acaggctcat gccggccgcc gccgcctttt
    10561 cctcaatcgc tcttcgttcg tctggaaggc agtacacctt gataggtggg ctgcccttcc
    10621 tggttggctt ggtttcatca gccatccgct tgccctcatc tgttacgccg gcggtagccg
    10681 gccagcctcg cagagcagga ttcccgttga gcaccgccag gtgcgaataa gggacagtga
    10741 agaaggaaca cccgctcgcg ggtgggccta cttcacctat cctgcccggc tgacgccgtt
    10801 ggatacacca aggaaagtct acacgaaccc tttggcaaaa tcctgtatat cgtgcgaaaa
    10861 aggatggata taccgaaaaa atcgctataa tgaccccgaa gcagggttat gcagcgga
//
