LOCUS       sequence-399551-       10593 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Frame -2 selector for NHEJ knock-in.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10593)
  AUTHORS   Shi Y, Kopparapu N, Ohler L, Dickinson DJ
  TITLE     Efficient and rapid fluorescent protein knock-in with universal 
            donors in mouse embryonic stem cells.
  JOURNAL   Development. 2023 May 15;150(10):dev201367. doi: 10.1242/dev.201367.
            Epub 2023 May 25.
  PUBMED    37129004
REFERENCE   2  (bases 1 to 10593)
  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: 
            "Development."; date: "2023-05-15"; volume: "150(10):dev201367. 
            doi"; pages: " 10.1242/dev.201367. Epub 2023 May 25"
COMMENT     Sequence Label: pYS19
FEATURES             Location/Qualifiers
     source          1..10593
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             8..477
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             946..1362
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1292..1314
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1330..1346
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1373..1621
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1373..1393
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1544..1563
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1643..1718
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        1771..2953
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2002..2944
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2901..2921
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      2956..2965
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2965..2988
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2989..3693
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(3131..3149)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3399..3418
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3718..3771
                     /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             3772..7875
                     /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             3772..7875
                     /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             7888..7908
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     LTR             8087..8556
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(8686..8703)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8857..9445)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8937..8956)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9605..10396,10397..10465))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     10228..10247
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10466..10570)
                     /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 caagctggcc 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 agcgctgcag catcgttctg
      901 tgttgtctct gtctgactgt gtttctgtat ttgtctgaaa attagggcca gactgttacc
      961 actcccttaa gtttgacctt aggtcactgg aaagatgtcg agcggatcgc tcacaaccag
     1021 tcggtagatg tcaagaagag acgttgggtt accttctgct ctgcagaatg gccaaccttt
     1081 aacgtcggat ggccgcgaga cggcaccttt aaccgagacc tcatcaccca ggttaagatc
     1141 aaggtctttt cacctggccc gcatggacac ccagaccagg tcccctacat cgtgacctgg
     1201 gaagccttgg cttttgaccc ccctccctgg gtcaagccct ttgtacaccc taagcctccg
     1261 cctcctcttc ctccatccgc cccgtctctc ccccttgaac ctcctcgttc gaccccgcct
     1321 cgatcctccc tttatccagc cctcactcct tctctaggcg ccggccggat ccgagggcct
     1381 atttcccatg attccttcat atttgcatat acgatacaag gctgttagag agataattgg
     1441 aattaatttg actgtaaaca caaagatatt agtacaaaat acgtgacgta gaaagtaata
     1501 atttcttggg tagtttgcag ttttaaaatt atgttttaaa atggactatc atatgcttac
     1561 cgtaacttga aagtatttcg atttcttggc tttatatatc ttgtggaaag gacgaaacac
     1621 cgcgggccag tacccaaaaa gcgttttaga gctagaaata gcaagttaaa ataaggctag
     1681 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt tttttcgaat tccgatgtac
     1741 gggccagata tacgcgttat catcgcgtga ggctccggtg cccgtcagtg ggcagagcgc
     1801 acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac cggtgcctag
     1861 agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc
     1921 gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac
     1981 gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc tggcctcttt
     2041 acgggttatg gcccttgcgt gccttgaatt acttccacgc ccctggctgc agtacgtgat
     2101 tcttgatccc gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg
     2161 agccccttcg cctcgtgctt gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg
     2221 aatctggtgg caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa
     2281 tttttgatga cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca
     2341 agatctgcac actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc
     2401 ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg
     2461 gtagtctcaa gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc
     2521 gccctgggcg gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct
     2581 tcccggccct gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg
     2641 tgagtcaccc acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc
     2701 cacggagtac cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc
     2761 gtctttaggt tggggggagg ggttttatgc gatggagttt ccccacactg agtgggtgga
     2821 gactgaagtt aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga
     2881 gtttggatct tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt
     2941 tcaggtgtcg tgagggccac catggattat aaggatgacg acgataaggt gagcaagggc
     3001 gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc
     3061 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
     3121 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc
     3181 ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc
     3241 gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag
     3301 gacggcggcg tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac
     3361 aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc
     3421 atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag
     3481 atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc
     3541 tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac
     3601 atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc
     3661 cactccaccg gcggcatgga cgagctgtac aaggtcgcta ccatcgaggg atccggagag
     3721 ggcagaggaa gtctgctaac atgcggtgac gtcgaggaga atcctggccc aatggacaag
     3781 aagtacagca tcggcctgga catcggcacc aacagcgtgg gctgggccgt gatcaccgac
     3841 gagtacaagg tgcccagcaa gaagttcaag gtgctgggca acaccgaccg ccacagcatc
     3901 aagaagaacc tgatcggcgc cctgctgttc gacagcggcg agaccgccga ggccacccgc
     3961 ctgaagcgca ccgcccgccg ccgctacacc cgccgcaaga accgcatctg ctacctgcag
     4021 gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccaccg cctggaggag
     4081 agcttcctgg tggaggagga caagaagcac gagcgccacc ccatcttcgg caacatcgtg
     4141 gacgaggtgg cctaccacga gaagtacccc accatctacc acctgcgcaa gaagctggtg
     4201 gacagcaccg acaaggccga cctgcgcctg atctacctgg ccctggccca catgatcaag
     4261 ttccgcggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga cgtggacaag
     4321 ctgttcatcc agctggtgca gacctacaac cagctgttcg aggagaaccc catcaacgcc
     4381 agcggcgtgg acgccaaggc catcctgagc gcccgcctga gcaagagccg ccgcctggag
     4441 aacctgatcg cccagctgcc cggcgagaag aagaacggcc tgttcggcaa cctgatcgcc
     4501 ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga ggacgccaag
     4561 ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc ccagatcggc
     4621 gaccagtacg ccgacctgtt cctggccgcc aagaacctga gcgacgccat cctgctgagc
     4681 gacatcctgc gcgtgaacac cgagatcacc aaggcccccc tgagcgccag catgatcaag
     4741 cgctacgacg agcaccacca ggacctgacc ctgctgaagg ccctggtgcg ccagcagctg
     4801 cccgagaagt acaaggagat cttcttcgac cagagcaaga acggctacgc cggctacatc
     4861 gacggcggcg ccagccagga ggagttctac aagttcatca agcccatcct ggagaagatg
     4921 gacggcaccg aggagctgct ggtgaagctg aaccgcgagg acctgctgcg caagcagcgc
     4981 accttcgaca acggcagcat cccccaccag atccacctgg gcgagctgca cgccatcctg
     5041 cgccgccagg aggacttcta ccccttcctg aaggacaacc gcgagaagat cgagaagatc
     5101 ctgaccttcc gcatccccta ctacgtcggc cccctggccc gcggcaacag ccgcttcgcc
     5161 tggatgaccc gcaagagcga ggagaccatc accccctgga acttcgagga ggtggtggac
     5221 aagggcgcca gcgcccagag cttcatcgag cgcatgacca acttcgacaa gaacctgccc
     5281 aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt gtacaacgag
     5341 ctgaccaagg tgaagtacgt gaccgagggc atgcgcaagc ccgccttcct gagcggcgag
     5401 cagaagaagg ccatcgtgga cctgctgttc aagaccaacc gcaaggtgac cgtgaagcag
     5461 ctgaaggagg actacttcaa gaagatcgag tgcttcgaca gcgtggagat cagcggcgtg
     5521 gaggaccgct tcaacgccag cctgggcacc taccacgacc tgctgaagat catcaaggac
     5581 aaggacttcc tggacaacga ggagaacgag gacatcctgg aggacatcgt gctgaccctg
     5641 accctgttcg aggaccgcga gatgatcgag gagcgcctga agacctacgc ccacctgttc
     5701 gacgacaagg tgatgaagca gctgaagcgc cgccgctaca ccggctgggg ccgcctgagc
     5761 cgcaagctga tcaacggcat ccgcgacaag cagagcggca agaccatcct ggacttcctg
     5821 aagagcgacg gcttcgccaa ccgcaacttc atgcagctga tccacgacga cagcctgacc
     5881 ttcaaggagg acatccagaa ggcccaggtg agcggccagg gcgacagcct gcacgagcac
     5941 atcgccaacc tggccggcag ccccgccatc aagaagggca tcctgcagac cgtgaaggtg
     6001 gtggacgagc tggtgaaggt gatgggccgc cacaagcccg agaacatcgt gatcgagatg
     6061 gcccgcgaga accagaccac ccagaagggc cagaagaaca gccgcgagcg catgaagcgc
     6121 atcgaggagg gcatcaagga gctgggcagc cagatcctga aggagcaccc cgtggagaac
     6181 acccagctgc agaacgagaa gctgtacctg tactacctgc agaacggccg cgacatgtac
     6241 gtggaccagg agctggacat caaccgcctg agcgactacg acgtggacca catcgtgccc
     6301 cagagcttcc tgaaggacga cagcatcgac aacaaggtgc tgacccgcag cgacaagaac
     6361 cgcggcaaga gcgacaacgt gcccagcgag gaggtggtga agaagatgaa gaactactgg
     6421 cgccagctgc tgaacgccaa gctgatcacc cagcgcaagt tcgacaacct gaccaaggcc
     6481 gagcgcggcg gcctgagcga gctggacaag gccggcttca tcaagcgcca gctggtggag
     6541 acccgccaga tcaccaagca cgtggcccag atcctggaca gccgcatgaa caccaagtac
     6601 gacgagaacg acaagctgat ccgcgaggtg aaggtgatca ccctgaagag caagctggtg
     6661 agcgacttcc gcaaggactt ccagttctac aaggtgcgcg agatcaacaa ctaccaccac
     6721 gcccacgacg cctacctgaa cgccgtggtg ggcaccgccc tgatcaagaa gtaccccaag
     6781 ctggagagcg agttcgtgta cggcgactac aaggtgtacg acgtgcgcaa gatgatcgcc
     6841 aagagcgagc aggagatcgg caaggccacc gccaagtact tcttctacag caacatcatg
     6901 aacttcttca agaccgagat caccctggcc aacggcgaga tccgcaagcg ccccctgatc
     6961 gagaccaacg gcgagaccgg cgagatcgtg tgggacaagg gccgcgactt cgccaccgtg
     7021 cgcaaggtgc tgagcatgcc ccaggtgaac atcgtgaaga agaccgaggt gcagaccggc
     7081 ggcttcagca aggagagcat cctgcccaag cgcaacagcg acaagctgat cgcccgcaag
     7141 aaggactggg accccaagaa gtacggcggc ttcgacagcc ccaccgtggc ctacagcgtg
     7201 ctggtggtgg ccaaggtgga gaagggcaag agcaagaagc tgaagagcgt gaaggagctg
     7261 ctgggcatca ccatcatgga gcgcagcagc ttcgagaaga accccatcga cttcctggag
     7321 gccaagggct acaaggaggt gaagaaggac ctgatcatca agctgcccaa gtacagcctg
     7381 ttcgagctgg agaacggccg caagcgcatg ctggccagcg ccggcgagct gcagaagggc
     7441 aacgagctgg ccctgcccag caagtacgtg aacttcctgt acctggccag ccactacgag
     7501 aagctgaagg gcagccccga ggacaacgag cagaagcagc tgttcgtgga gcagcacaag
     7561 cactacctgg acgagatcat cgagcagatc agcgagttca gcaagcgcgt gatcctggcc
     7621 gacgccaacc tggacaaggt gctgagcgcc tacaacaagc accgcgacaa gcccatccgc
     7681 gagcaggccg agaacatcat ccacctgttc accctgacca acctgggcgc ccccgccgcc
     7741 ttcaagtact tcgacaccac catcgaccgc aagcgctaca ccagcaccaa ggaggtgctg
     7801 gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacccgcat cgacctgagc
     7861 cagctgggcg gcgacagccg cgccgacccc aagaagaagc gcaaggtgtg agcggccgca
     7921 tcgataaaat aaaagatttt atttagtctc cagaaaaagg ggggaatgaa agaccccacc
     7981 tgtaggtttg gcaagctagc ttaagtaacg ccattttgca aggcatggaa aaatacataa
     8041 ctgagaatag agaagttcag atcaaggtca ggaacagatg gaacagctga atatgggcca
     8101 aacaggatat ctgtggtaag cagttcctgc cccggctcag ggccaagaac agatggaaca
     8161 gctgaatatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg ctcagggcca
     8221 agaacagatg gtccccagat gcggtccagc cctcagcagt ttctagagaa ccatcagatg
     8281 tttccagggt gccccaagga cctgaaatga ccctgtgcct tatttgaact aaccaatcag
     8341 ttcgcttctc gcttctgttc gcgcgcttct gctccccgag ctcaataaaa gagcccacaa
     8401 cccctcactc ggggcgccag tcctccgatt gactgagtcg cccgggtacc cgtgtatcca
     8461 ataaaccctc ttgcagttgc atccgacttg tggtctcgct gttccttggg agggtctcct
     8521 ctgagtgatt gactacccgt cagcgggggt ctttcacatg cagcatgtat caaaattaat
     8581 ttggtttttt ttcttaagta tttacattaa atggccatag ttgcattaat gaatcggcca
     8641 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     8701 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     8761 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     8821 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     8881 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     8941 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     9001 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     9061 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     9121 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     9181 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     9241 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     9301 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     9361 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     9421 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     9481 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     9541 cacctagatc cttttgcggc cgcaaatcaa tctaaagtat atatgagtaa acttggtctg
     9601 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     9661 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     9721 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     9781 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     9841 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     9901 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     9961 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
    10021 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
    10081 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
    10141 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
    10201 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
    10261 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
    10321 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
    10381 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
    10441 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
    10501 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
    10561 gggttccgcg cacatttccc cgaaaagtgc cac
//
