LOCUS       sequence-306195-       10063 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP-2A-Cas9 (CHO codon-optimized).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10063)
  AUTHORS   Sergeeva D, Lee GM, Nielsen LK, Grav LM
  TITLE     Multi-copy targeted integration for accelerated development of 
            high-producing CHO cells.
  JOURNAL   ACS Synth Biol. 2020 Aug 24. doi: 10.1021/acssynbio.0c00322.
  PUBMED    32835482
REFERENCE   2  (bases 1 to 10063)
  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: "ACS Synth 
            Biol. 2020 Aug 24. doi: 10.1021/acssynbio.0c00322."
COMMENT     Sequence Label: pJ607-CMV-GFP-2A-Cas9
FEATURES             Location/Qualifiers
     source          1..10063
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      86..674
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     575..594
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     828..845
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     943..960
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     944..960
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1416..1432
                     /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)."
     enhancer        1459..1838
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1839..2042
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1993..2013
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2087..2106
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2087..2105
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2129..2131,2132..2134,2135..2137,2138..2848)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MHVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLT
                     LKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKD
                     DGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGI
                     KVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVL
                     LEFVTAAGITLGMDELYK"
     primer_bind     complement(2177..2198)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2438..2457)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2785..2806
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2864..2917
                     /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             2930..7033
                     /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             2930..7033
                     /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             7046..7066
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(7078..7095)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7084..7195
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7313..7333)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7318..7647
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7498..7633
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7560..7579
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             7696..8721
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             7696..8721
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    8851..8972
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8888..8907)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8942..8961
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(9025..9816,9817..9885))
                     /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     9648..9667
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9886..9977)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     10019..10038
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     10044..10063
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 ctcatgacca aaatccctta acgtgagtta cgcgcgcgtc gttccactga gcgtcagacc
       61 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
      121 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
      181 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
      241 tgtagccgta gttagcccac cacttcaaga actctgtagc accgcctaca tacctcgctc
      301 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
      361 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
      421 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
      481 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
      541 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
      601 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
      661 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
      721 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
      781 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
      841 gcgaggaagc ggaaggcgag agtagggaac tgccaggcat caaactaagc agaaggcccc
      901 tgacggatgg cctttttgcg tttctacaaa ctctttctgt gttgtaaaac gacggccagt
      961 cttaagctcg ggccccctgg gcggttctga taacgagtaa tcgttaatcc gcaaataacg
     1021 taaaaacccg cttcggcggg tttttttatg gggggagttt agggaaagag catttgtcag
     1081 aatatttaag ggcgcctgtc actttgcttg atatatgaga attatttaac cttataaatg
     1141 agaaaaaagc aacgcacttt aaataagata cgttgctttt tcgattgatg aacacctata
     1201 attaaactat tcatctatta tttatgattt tttgtatata caatatttct agtttgttaa
     1261 agagaattaa gaaaataaat ctcgaaaata ataaagggaa aatcagtttt tgatatcaaa
     1321 attatacatg tcaacgataa tacaaaatat aatacaaact ataagatgtt atcagtattt
     1381 attatcattt agaataaatt ttgtgtcgcc cttaattgtg agcggataac aattacgagc
     1441 ttcatgcaca gtggcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
     1501 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     1561 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     1621 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     1681 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     1741 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     1801 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     1861 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     1921 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     1981 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     2041 ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat acgactcact
     2101 ataggggaag ctggctagcg ccgccaccat gcatgtgagc aagggcgagg agctgttcac
     2161 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     2221 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
     2281 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
     2341 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     2401 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     2461 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     2521 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     2581 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     2641 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     2701 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa
     2761 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     2821 cactctcggc atggacgagc tgtacaagcc taggggcagt ggagagggca gaggaagtct
     2881 gctaacatgc ggtgacgtcg aggagaatcc tggcccacga tcgagaagca tggacaagaa
     2941 atactccatc ggccttgata tcgggaccaa cagcgtgggt tgggccgtga tcactgacga
     3001 atacaaagtg ccatcgaaaa agtttaaggt gctgggcaat actgacagac atagcatcaa
     3061 gaaaaacctg attggagccc tcctgttcga ctccggggaa accgccgaag ccactcgcct
     3121 gaaacggacc gcacgccgga ggtacactcg ccggaagaat agaatctgct accttcaaga
     3181 gatctttagc aacgagatgg ccaaagtgga cgattcgttc ttccaccggc tggaggagtc
     3241 attcctggtc gaagaggaca aaaagcatga gagacatccg atcttcggta acattgtgga
     3301 tgaggtcgcc taccacgaga agtaccccac tatctaccat ttgcgcaaga agttggtcga
     3361 ctcgactgat aaggccgacc tcagactcat ctacctcgct ttggcacaca tgatcaagtt
     3421 taggggtcac ttcctgattg aaggggatct gaacccagac aactcggatg tggataaact
     3481 gtttatccag ctggtccaga cttacaatca gctctttgag gagaacccga tcaatgcctc
     3541 cggcgtggat gctaaggcaa tcctgtccgc acgcttgtca aagagccggc ggctcgaaaa
     3601 tctgatcgca cagcttccgg gagaaaagaa aaacggactg tttggaaacc ttatcgccct
     3661 ctcgctggga ctcactccta acttcaaatc caattttgat ttggccgagg atgccaagct
     3721 gcagctgtcg aaggacacct acgacgatga cctcgacaat cttctggccc agatcgggga
     3781 ccagtacgcc gatctgttcc tcgcggccaa aaacctgtca gacgcaatct tgctgagcga
     3841 tatcttgagg gtcaataccg aaatcactaa ggctccattg tcagcatcga tgatcaagag
     3901 gtacgatgaa caccatcagg acctgaccct gctcaaggca ctggtgcggc agcaactgcc
     3961 ggaaaagtac aaggagatct tcttcgatca aagcaagaat gggtacgcag ggtacatcga
     4021 tggtggagca tcacaagaag agttctacaa attcatcaag cccatccttg aaaagatgga
     4081 cgggacggaa gaactgctgg tgaagctgaa tcgggaagat ctgctgcgga agcagcgcac
     4141 cttcgacaat ggttccatcc cacaccaaat ccatctcggc gaactgcacg cgatccttcg
     4201 ccggcaggaa gatttctacc cgttcttgaa ggataacaga gaaaagatcg agaaaatcct
     4261 gacctttaga atcccgtact acgtgggccc gttggctcgc ggaaactcaa gattcgcctg
     4321 gatgactaga aaatccgaag aaacgattac cccgtggaac tttgaagagg tcgtcgacaa
     4381 gggagcctcg gcacagtcgt tcatcgaacg gatgaccaat ttcgacaaga acctccccaa
     4441 cgaaaaggtg ctcccgaaac actcgctgct gtatgagtac ttcacggtct acaacgagct
     4501 gaccaaggtg aagtacgtga ccgaaggaat gcggaagccc gcttttctgt ccggtgaaca
     4561 gaaaaaggcc atcgtggacc tcctcttcaa gactaacaga aaggtgaccg tgaaacagct
     4621 taaagaggac tacttcaaga agatcgagtg ttttgactcc gtggaaatct cgggagtgga
     4681 ggataggttt aacgcttcgc tggggaccta ccatgacctt ctcaagatca tcaaggataa
     4741 ggatttcctg gacaacgaag aaaacgagga catccttgag gacatcgtgt tgaccttgac
     4801 cctgtttgag gatcgggaaa tgatcgagga gagactgaaa acttacgcgc atcttttcga
     4861 cgacaaagtg atgaagcagc tgaaaagaag aagatacact ggctggggac ggctgtcgag
     4921 gaaactgatt aacggaatta gggataaaca aagcggaaag actatcctcg attttctcaa
     4981 gtcggacggc ttcgccaatc ggaactttat gcagctcatc cacgacgata gcctgacctt
     5041 caaggaagat atccaaaaag cccaagtgtc cggccaggga gatagcttgc acgagcacat
     5101 tgctaacctg gccggttcac cagccattaa gaagggaatc ctccaaaccg tgaaggtggt
     5161 cgacgaactc gtcaaggtga tgggccgcca taagccagag aacatcgtga tcgagatggc
     5221 tcgcgaaaat cagactactc agaagggtca aaagaactcc cgcgagcgca tgaagcgcat
     5281 cgaagaagga attaaggagc tgggctcaca aatcctcaaa gaacatcctg tcgagaacac
     5341 ccaacttcag aacgagaaac tgtacctcta ctatctccaa aatggccggg acatgtacgt
     5401 cgatcaggaa ttggatatca accgcctctc agactacgac gtggatcata tcgtgccgca
     5461 gtcgtttctc aaagatgata gcatcgacaa caaggtcctg acgaggtcgg ataagaaccg
     5521 cgggaaatcc gacaatgtgc cttcggaaga ggtggtgaaa aagatgaaga actactggag
     5581 gcaattgctt aatgccaaac tgatcaccca gcgcaaattc gacaacctta ctaaggccga
     5641 gcgcggagga ctgtccgaac tcgataaggc ggggttcatc aaaagacaat tggtcgaaac
     5701 tcggcaaatt accaagcatg tcgctcagat cctcgactcg cgcatgaaca ctaagtatga
     5761 tgagaacgac aagctcattc gcgaagtgaa agtgattacc ctcaaatcaa agctggtgtc
     5821 ggacttcaga aaagactttc aattctacaa ggtgcgggag attaacaact accaccacgc
     5881 gcacgacgcc tacctcaatg cagtggtggg aaccgccctc atcaagaagt atccaaagct
     5941 cgaaagcgag ttcgtgtacg gggactataa agtgtacgat gtgcggaaaa tgatcgcaaa
     6001 gagcgagcaa gagatcggca aagcaacggc caaatacttc ttctacagca acattatgaa
     6061 ctttttcaag accgagatta ccctggcgaa cggagaaatc cggaagcggc ctttgatcga
     6121 aacgaatggg gaaactggag aaatcgtgtg ggacaagggc agagactttg cgaccgtgag
     6181 aaaggtgctg tccatgccac aagtcaacat cgtgaagaaa actgaagtgc agactggagg
     6241 attttccaag gagtcaatcc ttccgaagcg caacagcgac aagctgatcg ccaggaagaa
     6301 ggattgggac cccaagaagt acggtggttt tgattcacct actgtcgctt acagcgtgct
     6361 ggtggtggcc aaggtggaaa aggggaagtc aaagaaattg aagtcggtga aggaattgct
     6421 ggggattact atcatggaga ggagcagctt cgaaaagaat cccatcgact tcttggaggc
     6481 caagggatac aaagaggtga agaaagacct gatcattaag ctgccgaagt actccttgtt
     6541 cgaactggaa aacggcagaa agcggatgct ggcctccgcc ggagagctgc agaagggtaa
     6601 cgagctggct ctgcccagca aatacgtgaa tttcctctac ctggcctccc attacgagaa
     6661 gctcaaggga agcccggagg ataatgagca aaaacaactg ttcgtcgagc agcacaagca
     6721 ctacctcgac gagatcatcg aacaaatctc cgagttctcg aagcgggtga ttctggccga
     6781 cgcaaacctt gataaagtcc tcagcgccta caacaagcac cgcgacaaac cgatcagaga
     6841 acaagctgag aacatcatcc acctgttcac gctgactaac ttgggagcgc ctgccgcctt
     6901 caagtacttc gacaccacta tcgatcggaa acggtacact tccactaagg aagtgctgga
     6961 cgcaaccctg atccatcagt ccatcaccgg actttatgag actcggatcg atctcagcca
     7021 gcttggcggt gattcaagag ctgacccgaa gaagaagcgc aaggtctaga aaatcagcct
     7081 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     7141 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcacaac
     7201 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     7261 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     7321 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     7381 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     7441 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     7501 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     7561 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     7621 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     7681 tctgatcagc acgtgatgaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg
     7741 atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt
     7801 gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat
     7861 ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg
     7921 gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca
     7981 cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc
     8041 gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca
     8101 ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct
     8161 gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg
     8221 caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg
     8281 cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt
     8341 gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg
     8401 aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag
     8461 cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat
     8521 cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca
     8581 atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc
     8641 gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact
     8701 cgtccgaggg caaaggaata gcacgtgcta cgagatttcg attccaccgc cgccttctat
     8761 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     8821 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     8881 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     8941 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     9001 tagagcttgg cgtaatcatg gtcattacca atgcttaatc agtgaggcac ctatctcagc
     9061 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     9121 acgggagggc ttaccatctg gccccagcgc tgcgatgata ccgcgagaac cacgctcacc
     9181 ggctccggat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     9241 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     9301 ttcgccagtt aatagtttgc gcaacgttgt tgccatcgct acaggcatcg tggtgtcacg
     9361 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     9421 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     9481 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     9541 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     9601 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     9661 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     9721 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     9781 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     9841 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatattct tcctttttca
     9901 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     9961 ttagaaaaat aaacaaatag gggtcagtgt tacaaccaat taaccaattc tgaacattat
    10021 cgcgagccca tttatacctg aatatggctc ataacacccc ttg
//
