LOCUS       sequence-153107-       10048 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cas9, mCherry, and sgRNA expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10048)
  AUTHORS   Schmid-Burgk JL, Honing K, Ebert TS, Hornung V
  TITLE     CRISPaint allows modular base-specific gene tagging using a 
            ligase-4-dependent mechanism.
  JOURNAL   Nat Commun. 2016 Jul 28;7:12338. doi: 10.1038/ncomms12338.
  PUBMED    27465542
REFERENCE   2  (bases 1 to 10048)
  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: "2016-07-28"; volume: "7:12338. doi"; pages: " 
            10.1038/ncomms12338"
COMMENT     Sequence Label: pCAS9-mCherry empty
FEATURES             Location/Qualifiers
     source          1..10048
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             8..477
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             938..1354
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1322..1338
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1365..1613
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1365..1385
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1536..1555
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        1712..2091
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2092..2295
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2246..2266
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2350..3057
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2495..2513)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2763..2782
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3082..3135
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3136..7239
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3136..7239
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7252..7272
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     LTR             7329..7922
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(8123..8140)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8294..8882)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8374..8393)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9060..9851,9852..9920))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     9683..9702
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9921..10025)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgcagcctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
       61 gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt
      121 tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag ccctcagcag
      181 tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg accctgtgcc
      241 ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga
      301 gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat tgactgagtc
      361 gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc
      421 tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcattt
      481 gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac caccgggagg
      541 taagctggcc agcaacttat ctgtgtctgt ccgattgtct agtgtctatg actgatttta
      601 tgcgcctgcg tcggtactag ctagttagct aactagctct gtatctggcg gacccgtggt
      661 ggaactgacg agttctgaac acccggccgc aaccctggga gacgtcccag ggactttggg
      721 ggccgttttt gtggcccgac ctgaggaagg gagtcgatgt ggaatccgac cccgtcagga
      781 tatgtggttc tggtaggaga cgagaaccta aaacagttcc cgcctccgtc tgaatttttg
      841 ctttcggttt ggaaccgaag ccgcgcgtct tgtctgctgc agcatcgttc tgtgttgtct
      901 ctgtctgact gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt
      961 aagtttgacc ttagatcact ggaaagatgt cgagcggctc gctcacaacc agtcggtaga
     1021 tgtcaagaag agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg
     1081 atggccgcga gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt
     1141 ttcacctggc ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt
     1201 ggcttttgac ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct
     1261 tcttccatcc gcgccgtctc tcccccttga acctcctctt tcgaccccgc ctcaatcctc
     1321 cctttatcca gccctcactc cttctctagg cgccggccgg atccgagggc ctatttccca
     1381 tgattccttc atatttgcat atacgataca aggctgttag agagataatt ggaattaatt
     1441 tgactgtaaa cacaaagata ttagtacaaa atacgtgacg tagaaagtaa taatttcttg
     1501 ggtagtttgc agttttaaaa ttatgtttta aaatggacta tcatatgctt accgtaactt
     1561 gaaagtattt cgatttcttg gctttatata tcttgtggaa aggacgaaac accgggccct
     1621 cgatatcgac tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg ctttttttcg
     1681 aattccgatg tacgggccag atatacgcgt tgacattgat tattgtctag ttattaatag
     1741 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     1801 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     1861 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     1921 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
     1981 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
     2041 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
     2101 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
     2161 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
     2221 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
     2281 tatataagca gagctctctg gctaactaga gaacccactg cttactggct tatcgaaatt
     2341 aagcttccca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg
     2401 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
     2461 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc
     2521 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac
     2581 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag
     2641 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     2701 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac
     2761 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc
     2821 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac
     2881 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc
     2941 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa
     3001 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaaggtc
     3061 gctaccatcg agggatccgg agagggcaga ggaagtctgc taacatgcgg tgacgtcgag
     3121 gagaatcctg gcccaatgga caagaagtac agcatcggcc tggacatcgg caccaacagc
     3181 gtgggctggg ccgtgatcac cgacgagtac aaggtgccca gcaagaagtt caaggtgctg
     3241 ggcaacaccg accgccacag catcaagaag aacctgatcg gcgccctgct gttcgacagc
     3301 ggcgagaccg ccgaggccac ccgcctgaag cgcaccgccc gccgccgcta cacccgccgc
     3361 aagaaccgca tctgctacct gcaggagatc ttcagcaacg agatggccaa ggtggacgac
     3421 agcttcttcc accgcctgga ggagagcttc ctggtggagg aggacaagaa gcacgagcgc
     3481 caccccatct tcggcaacat cgtggacgag gtggcctacc acgagaagta ccccaccatc
     3541 taccacctgc gcaagaagct ggtggacagc accgacaagg ccgacctgcg cctgatctac
     3601 ctggccctgg cccacatgat caagttccgc ggccacttcc tgatcgaggg cgacctgaac
     3661 cccgacaaca gcgacgtgga caagctgttc atccagctgg tgcagaccta caaccagctg
     3721 ttcgaggaga accccatcaa cgccagcggc gtggacgcca aggccatcct gagcgcccgc
     3781 ctgagcaaga gccgccgcct ggagaacctg atcgcccagc tgcccggcga gaagaagaac
     3841 ggcctgttcg gcaacctgat cgccctgagc ctgggcctga cccccaactt caagagcaac
     3901 ttcgacctgg ccgaggacgc caagctgcag ctgagcaagg acacctacga cgacgacctg
     3961 gacaacctgc tggcccagat cggcgaccag tacgccgacc tgttcctggc cgccaagaac
     4021 ctgagcgacg ccatcctgct gagcgacatc ctgcgcgtga acaccgagat caccaaggcc
     4081 cccctgagcg ccagcatgat caagcgctac gacgagcacc accaggacct gaccctgctg
     4141 aaggccctgg tgcgccagca gctgcccgag aagtacaagg agatcttctt cgaccagagc
     4201 aagaacggct acgccggcta catcgacggc ggcgccagcc aggaggagtt ctacaagttc
     4261 atcaagccca tcctggagaa gatggacggc accgaggagc tgctggtgaa gctgaaccgc
     4321 gaggacctgc tgcgcaagca gcgcaccttc gacaacggca gcatccccca ccagatccac
     4381 ctgggcgagc tgcacgccat cctgcgccgc caggaggact tctacccctt cctgaaggac
     4441 aaccgcgaga agatcgagaa gatcctgacc ttccgcatcc cctactacgt cggccccctg
     4501 gcccgcggca acagccgctt cgcctggatg acccgcaaga gcgaggagac catcaccccc
     4561 tggaacttcg aggaggtggt ggacaagggc gccagcgccc agagcttcat cgagcgcatg
     4621 accaacttcg acaagaacct gcccaacgag aaggtgctgc ccaagcacag cctgctgtac
     4681 gagtacttca ccgtgtacaa cgagctgacc aaggtgaagt acgtgaccga gggcatgcgc
     4741 aagcccgcct tcctgagcgg cgagcagaag aaggccatcg tggacctgct gttcaagacc
     4801 aaccgcaagg tgaccgtgaa gcagctgaag gaggactact tcaagaagat cgagtgcttc
     4861 gacagcgtgg agatcagcgg cgtggaggac cgcttcaacg ccagcctggg cacctaccac
     4921 gacctgctga agatcatcaa ggacaaggac ttcctggaca acgaggagaa cgaggacatc
     4981 ctggaggaca tcgtgctgac cctgaccctg ttcgaggacc gcgagatgat cgaggagcgc
     5041 ctgaagacct acgcccacct gttcgacgac aaggtgatga agcagctgaa gcgccgccgc
     5101 tacaccggct ggggccgcct gagccgcaag ctgatcaacg gcatccgcga caagcagagc
     5161 ggcaagacca tcctggactt cctgaagagc gacggcttcg ccaaccgcaa cttcatgcag
     5221 ctgatccacg acgacagcct gaccttcaag gaggacatcc agaaggccca ggtgagcggc
     5281 cagggcgaca gcctgcacga gcacatcgcc aacctggccg gcagccccgc catcaagaag
     5341 ggcatcctgc agaccgtgaa ggtggtggac gagctggtga aggtgatggg ccgccacaag
     5401 cccgagaaca tcgtgatcga gatggcccgc gagaaccaga ccacccagaa gggccagaag
     5461 aacagccgcg agcgcatgaa gcgcatcgag gagggcatca aggagctggg cagccagatc
     5521 ctgaaggagc accccgtgga gaacacccag ctgcagaacg agaagctgta cctgtactac
     5581 ctgcagaacg gccgcgacat gtacgtggac caggagctgg acatcaaccg cctgagcgac
     5641 tacgacgtgg accacatcgt gccccagagc ttcctgaagg acgacagcat cgacaacaag
     5701 gtgctgaccc gcagcgacaa gaaccgcggc aagagcgaca acgtgcccag cgaggaggtg
     5761 gtgaagaaga tgaagaacta ctggcgccag ctgctgaacg ccaagctgat cacccagcgc
     5821 aagttcgaca acctgaccaa ggccgagcgc ggcggcctga gcgagctgga caaggccggc
     5881 ttcatcaagc gccagctggt ggagacccgc cagatcacca agcacgtggc ccagatcctg
     5941 gacagccgca tgaacaccaa gtacgacgag aacgacaagc tgatccgcga ggtgaaggtg
     6001 atcaccctga agagcaagct ggtgagcgac ttccgcaagg acttccagtt ctacaaggtg
     6061 cgcgagatca acaactacca ccacgcccac gacgcctacc tgaacgccgt ggtgggcacc
     6121 gccctgatca agaagtaccc caagctggag agcgagttcg tgtacggcga ctacaaggtg
     6181 tacgacgtgc gcaagatgat cgccaagagc gagcaggaga tcggcaaggc caccgccaag
     6241 tacttcttct acagcaacat catgaacttc ttcaagaccg agatcaccct ggccaacggc
     6301 gagatccgca agcgccccct gatcgagacc aacggcgaga ccggcgagat cgtgtgggac
     6361 aagggccgcg acttcgccac cgtgcgcaag gtgctgagca tgccccaggt gaacatcgtg
     6421 aagaagaccg aggtgcagac cggcggcttc agcaaggaga gcatcctgcc caagcgcaac
     6481 agcgacaagc tgatcgcccg caagaaggac tgggacccca agaagtacgg cggcttcgac
     6541 agccccaccg tggcctacag cgtgctggtg gtggccaagg tggagaaggg caagagcaag
     6601 aagctgaaga gcgtgaagga gctgctgggc atcaccatca tggagcgcag cagcttcgag
     6661 aagaacccca tcgacttcct ggaggccaag ggctacaagg aggtgaagaa ggacctgatc
     6721 atcaagctgc ccaagtacag cctgttcgag ctggagaacg gccgcaagcg catgctggcc
     6781 agcgccggcg agctgcagaa gggcaacgag ctggccctgc ccagcaagta cgtgaacttc
     6841 ctgtacctgg ccagccacta cgagaagctg aagggcagcc ccgaggacaa cgagcagaag
     6901 cagctgttcg tggagcagca caagcactac ctggacgaga tcatcgagca gatcagcgag
     6961 ttcagcaagc gcgtgatcct ggccgacgcc aacctggaca aggtgctgag cgcctacaac
     7021 aagcaccgcg acaagcccat ccgcgagcag gccgagaaca tcatccacct gttcaccctg
     7081 accaacctgg gcgcccccgc cgccttcaag tacttcgaca ccaccatcga ccgcaagcgc
     7141 tacaccagca ccaaggaggt gctggacgcc accctgatcc accagagcat caccggcctg
     7201 tacgagaccc gcatcgacct gagccagctg ggcggcgaca gccgcgccga ccccaagaag
     7261 aagcgcaagg tgtgagcggc cgcatcgata aaataaaaga ttttatttag tctccagaaa
     7321 aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt
     7381 tgcaaggcat ggaaaaatac ataactgaga atagagaagt tcagatcaag gtcaggaaca
     7441 gatggaacag ctgaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc
     7501 tcagggccaa gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc
     7561 agttcctgcc ccggctcagg gccaagaaca gatggtcccc agatgcggtc cagccctcag
     7621 cagtttctag agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt
     7681 gccttatttg aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc
     7741 cgagctcaat aaaagagccc acaacccctc actcggggcg ccagtcctcc gattgactga
     7801 gtcgcccggg tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct
     7861 cgctgttcct tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca
     7921 tttccgactt gtggtctcgc tgccttggga gggtctcctc tgagtgattg actacccgtc
     7981 agcgggggtc ttcacatgca gcatgtatca aaattaattt ggtttttttt cttaagtatt
     8041 tacattaaat ggccatagtt gcattaatga atcggccaac gcgcggggag aggcggtttg
     8101 cgtattggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     8161 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     8221 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     8281 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     8341 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     8401 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     8461 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     8521 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     8581 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     8641 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     8701 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     8761 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     8821 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     8881 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     8941 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     9001 aatgaagttt gcggccgcaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     9061 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     9121 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     9181 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     9241 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     9301 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     9361 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     9421 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     9481 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     9541 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     9601 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     9661 tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     9721 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     9781 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     9841 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     9901 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     9961 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
    10021 ccgcgcacat ttccccgaaa agtgccac
//
