LOCUS       sequence-222816-       12895 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dox-inducible Cas9 expression with a CD4 selection marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12895)
  AUTHORS   Hulton CH, Costa EA, Shah NS, Quintanal-Villalonga A, Heller G, de 
            Stanchina E, Rudin CM, Poirier JT
  TITLE     Direct genome editing of patient-derived xenografts using 
            CRISPR-Cas9 enables rapid in vivo functional genomics.
  JOURNAL   Nat Cancer. 2020 Mar;1(3):359-369. doi: 10.1038/s43018-020-0040-8. 
            Epub 2020 Mar 9.
  PUBMED    33345196
REFERENCE   2  (bases 1 to 12895)
  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"; doi: 
            "10.1038/s43018-020-0040-8"; journalName: "Nat Cancer"; date: 
            "2020-03"; volume: "1"; issue: "3"; pages: "359-369"
COMMENT     Sequence Label: pSpCTRE-CD4
FEATURES             Location/Qualifiers
     source          1..12895
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2028..2143
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     polyA_signal    2187..2321
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(2505..2528)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(2529..2576)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(2577..6680)
                     /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             complement(2577..6680)
                     /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"
     promoter        complement(6687..7054)
                     /label=TRE3GS promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, modified to eliminate
                     binding sites for endogenous mammalian transcription 
                     factors"
     protein_bind    6812..6830
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6848..6866
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6884..6902
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6920..6938
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6956..6974
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6992..7010
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7028..7046
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        7114..7325
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             8049..8102
                     /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             8103..8849
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     misc_feature    8863..9451
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8916..8936)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9334..9345)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9659..10292
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10420..10436)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10420..10436)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10433..10455)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10444..10460
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(10468..10474,10475..10492,10493..10498))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10513..10534
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10651..10668)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(10822..11410)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(10902..10921)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11581..12372,12373..12441))
                     /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     12204..12223
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12442..12546)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    12594..12728
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag
     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2101 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2161 acagcagaga tccagtttat cgacttaact tgtttattgc agcttataat ggttacaaat
     2221 aaggcaatag catcacaaat ttcacaaata aggcattttt ttcactgcat tctagttttg
     2281 gtttgtccaa actcatcaat gtatcttatc atgtctggat ctcaaatccc tcggaagctg
     2341 cgcctgtctt aggttggagt gatacatttt tatcactttt acccgtcttt ggattaggca
     2401 gtagctctga cggccctcct gtcttaggtt agtgaaaaat gtcactctct tacccgtcat
     2461 tggctgtcca gcttagctcg caggggaggt ggtctggatc cttacttatc gtcatcgtct
     2521 ttgtaatctt tcttcttctt agcctgtcca gccttctttg tggcggcagg tcgcttgtcg
     2581 cctcccagct gagacaggtc gatccgtgtc tcgtacaggc cggtgatgct ctggtggatc
     2641 agggtggcgt ccagcacctc tttggtgctg gtgtacctct tccggtcgat ggtggtgtca
     2701 aagtacttga aggcggcagg ggctcccaga ttggtcaggg taaacaggtg gatgatattc
     2761 tcggcctgct ctctgatggg cttatcccgg tgcttgttgt aggcggacag cactttgtcc
     2821 agattagcgt cggccaggat cactctcttg gagaactcgc tgatctgctc gatgatctcg
     2881 tccaggtagt gcttgtgctg ttccacaaac agctgtttct gctcattatc ctcgggggag
     2941 cccttcagct tctcatagtg gctggccagg tacaggaagt tcacatattt ggagggcagg
     3001 gccagttcgt ttcccttctg cagttcgccg gcagaggcca gcattctctt ccggccgttt
     3061 tccagctcga acagggagta cttaggcagc ttgatgatca ggtccttttt cacttctttg
     3121 tagcccttgg cttccagaaa gtcgatggga ttcttctcga agctgcttct ttccatgatg
     3181 gtgatcccca gcagctcttt cacactcttc agtttcttgg acttgccctt ttccactttg
     3241 gccaccacca gcacagaata ggccacggtg gggctgtcga agccgccgta cttcttaggg
     3301 tcccagtcct tctttctggc gatcagctta tcgctgttcc tcttgggcag gatagactct
     3361 ttgctgaagc cgcctgtctg cacctcggtc tttttcacga tattcacttg gggcatgctc
     3421 agcactttcc gcacggtggc aaaatcccgg cccttatccc acacgatctc cccggtttcg
     3481 ccgtttgtct cgatcagagg ccgcttccgg atctcgccgt tggccagggt aatctcggtc
     3541 ttgaaaaagt tcatgatgtt gctgtagaag aagtacttgg cggtagcctt gccgatttcc
     3601 tgctcgctct tggcgatcat cttccgcacg tcgtacacct tgtagtcgcc gtacacgaac
     3661 tcgctttcca gcttagggta ctttttgatc agggcggttc ccacgacggc gttcaggtag
     3721 gcgtcgtggg cgtggtggta gttgttgatc tcgcgcactt tgtaaaactg gaaatccttc
     3781 cggaaatcgg acaccagctt ggacttcagg gtgatcactt tcacttcccg gatcagcttg
     3841 tcattctcgt cgtacttagt gttcatccgg gagtccagga tctgtgccac gtgctttgtg
     3901 atctgccggg tttccaccag ctgtctcttg atgaagccgg ccttatccag ttcgctcagg
     3961 ccgcctctct cggccttggt cagattgtcg aactttctct gggtaatcag cttggcgttc
     4021 agcagctgcc gccagtagtt cttcatcttc ttcacgacct cttcggaggg cacgttgtcg
     4081 ctcttgcccc ggttcttgtc gcttctggtc agcaccttgt tgtcgatgga gtcgtccttc
     4141 agaaagctct gaggcacgat atggtccaca tcgtagtcgg acagccggtt gatgtccagt
     4201 tcctggtcca cgtacatatc ccgcccattc tgcaggtagt acaggtacag cttctcgttc
     4261 tgcagctggg tgttttccac ggggtgttct ttcaggatct ggctgcccag ctctttgatg
     4321 ccctcttcga tccgcttcat tctctcgcgg ctgttcttct gtcccttctg ggtggtctgg
     4381 ttctctctgg ccatttcgat cacgatgttc tcgggcttgt gccggcccat cactttcacg
     4441 agctcgtcca ccaccttcac tgtctgcagg atgcccttct taatggcggg gctgccggcc
     4501 agattggcaa tgtgctcgtg caggctatcg ccctggccgg acacctgggc tttctggatg
     4561 tcctctttaa aggtcaggct gtcgtcgtgg atcagctgca tgaagtttct gttggcgaag
     4621 ccgtcggact tcaggaaatc caggattgtc ttgccggact gcttgtcccg gatgccgttg
     4681 atcagcttcc ggctcagcct gccccagccg gtgtatctcc gccgcttcag ctgcttcatc
     4741 actttgtcgt cgaacaggtg ggcataggtt ttcagccgtt cctcgatcat ctctctgtcc
     4801 tcaaacagtg tcagggtcag cacgatatct tccagaatgt cctcgttttc ctcattgtcc
     4861 aggaagtcct tgtccttgat aattttcagc agatcgtggt atgtgcccag ggaggcgttg
     4921 aaccgatctt ccacgccgga gatttccacg gagtcgaagc actcgatttt cttgaagtag
     4981 tcctctttca gctgcttcac ggtcactttc cggttggtct tgaacagcag gtccacgatg
     5041 gcctttttct gctcgccgct caggaaggcg ggctttctca ttccctcggt cacgtatttc
     5101 actttggtca gctcgttata cacggtgaag tactcgtaca gcaggctgtg cttgggcagc
     5161 accttctcgt tgggcaggtt cttatcgaag ttggtcatcc gctcgatgaa gctctgggcg
     5221 gaagcgccct tgtccaccac ttcctcgaag ttccaggggg tgatggtttc ctcgctcttt
     5281 ctggtcatcc aggcgaatct gctgtttccc ctggccagag ggcccacgta gtaggggatg
     5341 cggaaggtca ggatcttctc gatcttttcc cggttgtcct tcaggaatgg gtaaaaatct
     5401 tcctgccgcc gcagaatggc gtgcagctct cccaggtgga tctggtgggg gatgctgccg
     5461 ttgtcgaagg tccgctgctt ccgcagcagg tcctctctgt tcagcttcac gagcagttcc
     5521 tcggtgccgt ccatcttttc caggatgggc ttgatgaact tgtagaactc ttcctggctg
     5581 gctccgccgt caatgtagcc ggcgtagccg ttcttgctct ggtcgaagaa aatctctttg
     5641 tacttctcag gcagctgctg ccgcacgaga gctttcagca gggtcaggtc ctggtggtgc
     5701 tcgtcgtatc tcttgatcat agaggcgctc aggggggcct tggtgatctc ggtgttcact
     5761 ctcaggatgt cgctcagcag gatggcgtcg gacaggttct tggcggccag aaacaggtcg
     5821 gcgtactggt cgccgatctg ggccagcagg ttgtccaggt cgtcgtcgta ggtgtccttg
     5881 ctcagctgca gtttggcatc ctcggccagg tcgaagttgc tcttgaagtt gggggtcagg
     5941 cccaggctca gggcaatcag gtttccgaac aggccattct tcttctcgcc gggcagctgg
     6001 gcgatcagat tttccagccg tctgctcttg ctcagtctgg cagacaggat ggccttggcg
     6061 tccacgccgc tggcgttgat ggggttttcc tcgaacagct ggttgtaggt ctgcaccagc
     6121 tggatgaaca gcttgtccac gtcgctgttg tcggggttca ggtcgccctc gatcaggaag
     6181 tggccccgga acttgatcat gtgggccagg gccagataga tcagccgcag gtcggccttg
     6241 tcggtgctgt ccaccagttt ctttctcagg tggtagatgg tggggtactt ctcgtggtag
     6301 gccacctcgt ccacgatgtt gccgaagatg gggtgccgct cgtgcttctt atcctcttcc
     6361 accaggaagg actcttccag tctgtggaag aagctgtcgt ccaccttggc catctcgttg
     6421 ctgaagatct cttgcagata gcagatccgg ttcttccgtc tggtgtatct tcttctggcg
     6481 gttctcttca gccgggtggc ctcggctgtt tcgccgctgt cgaacagcag ggctccgatc
     6541 aggttcttct tgatgctgtg ccggtcggtg ttgcccagca ccttgaattt cttgctgggc
     6601 accttgtact cgtcggtgat cacggcccag cccacagagt tggtgccgat gtccaggccg
     6661 atgctgtact tcttgtccat gaattcttta cgagggtagg aagtggtacg gaaagttggt
     6721 ataagacaaa agtgttgtgg aattgaagtt tactcaaaaa atcagcactc ttttataggc
     6781 gccctggttt acataagcaa agcttatacg ttctctatca ctgataggga gtaaactgga
     6841 tatacgttct ctatcactga tagggagtaa actgtagata cgttctctat cactgatagg
     6901 gagtaaactg gtcatacgtt ctctatcact gatagggagt aaactcctta tacgttctct
     6961 atcactgata gggagtaaag tctgcatacg ttctctatca ctgataggga gtaaactctt
     7021 catacgttct ctatcactga tagggagtaa actcgaggtg ataattccac taggtcttga
     7081 aaggagtggg aattggctcc ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc
     7141 ccgagaagtt ggggggaggg gtcggcaatt gatccggtgc ctagagaagg tggcgcgggg
     7201 taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt gggggagaac
     7261 cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt gccgccagaa
     7321 cacaggatcg atcacgtacg gccaccatga accggggagt cccttttagg cacttgcttc
     7381 tggtgctgca actggcgctc ctcccagcag ccactcaggg aaagaaagtg gtgctgggca
     7441 aaaaagggga tacagtggaa ctgacctgta cagcttccca gaagaagagc atacaattcc
     7501 actggaaaaa ctccaaccag ataaagattc tgggaaatca gggctccttc ttaactaaag
     7561 gtccatccaa gctgaatgat cgcgctgact caagaagaag cctttgggac caaggaaact
     7621 ttcccctgat catcaagaat cttaagatag aagactcaga tacttacatc tgtgaagtgg
     7681 aggaccagaa ggaggaggtg caattgctag tgttcggatt gactgccaac tctgacaccc
     7741 acctgcttca ggggcagagc ctgaccctga ccttggagag cccccctggt agtagcccct
     7801 cagtgcaatg taggagtcca aggggtaaaa acatacaggg ggggaagacc ctctccgtgt
     7861 ctcagctgga gctccaggat agtggcacct ggacatgcac tgtcttgcag aaccagaaga
     7921 aggtggagtt caaaatagac atcgtggtgc tagcttctgg cggtggctcc ggaatggccc
     7981 tgattgtgct ggggggcgtc gccggcctcc tgcttttcat tgggctaggc atcttcttcg
     8041 gctccggaga aggtagaggt tctctcctca cttgtggtga tgttgaagaa aaccctggtc
     8101 caatgtctag actggacaag agcaaagtca taaactctgc tctggaatta ctcaatggag
     8161 tcggtatcga aggcctgacg acaaggaaac tcgctcaaaa gctgggagtt gagcagccta
     8221 ccctgtactg gcacgtgaag aacaagcggg ccctgctcga tgccctgcca atcgagatgc
     8281 tggacaggca tcatacccac tcctgccccc tggaaggcga gtcatggcaa gactttctgc
     8341 ggaacaacgc caagtcatac cgctgtgctc tcctctcaca tcgcgacggg gctaaagtgc
     8401 atctcggcac ccgcccaaca gagaaacagt acgaaaccct ggaaaatcag ctcgcgttcc
     8461 tgtgtcagca aggcttctcc ctggagaacg cactgtacgc tctgtccgcc gtgggccact
     8521 ttacactggg ctgcgtattg gaggaacagg agcatcaagt agcaaaagag gaaagagaga
     8581 cacctaccac cgattctatg cccccacttc tgaaacaagc aattgagctg ttcgaccggc
     8641 agggagccga acctgccttc cttttcggcc tggaactaat catatgtggc ctggagaaac
     8701 agctaaagtg cgaaagcggc gggccgaccg acgcccttga cgattttgac ttagacatgc
     8761 tcccagccga tgcccttgac gactttgacc ttgatatgct gcctgctgac gctcttgacg
     8821 attttgacct tgacatgctc cccgggtaaa cgcgtctgga acaatcaacc tctggattac
     8881 aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga
     8941 tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc
     9001 tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa
     9061 cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc
     9121 acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc
     9181 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     9241 gtggtgttgt cggggaagct gacgtccttt ccatggctgc tcgcctgtgt tgccacctgg
     9301 attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct
     9361 tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg
     9421 agtcggatct ccctttgggc cgcctccccg cctggaatta attctgcagt cgagacctag
     9481 aaaaacatgg agcaatcaca agtagcaata cagcagctac caatgctgat tgtgcctggc
     9541 tagaagcaca agaggaggag gaggtgggtt tttccagtca cacctcaggt acctttaaga
     9601 ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa gaggggactg
     9661 gaagggctaa ttcactccca acgaagacaa gatatccttg atctgtggat ctaccacaca
     9721 caaggctact tccctgatta gcagaactac acaccagggc caggggtcag atatccactg
     9781 acctttggat ggtgctacaa gctagtacca gttgagccag ataaggtaga agaggccaat
     9841 aaaggagaga acaccagctt gttacaccct gtgagcctgc atgggatgga tgacccggag
     9901 agagaagtgt tagagtggag gtttgacagc cgcctagcat ttcatcacgt ggcccgagag
     9961 ctgcatccgg agtacttcaa gaactgctga tatcgagctt gctacaaggg actttccgct
    10021 ggggactttc cagggaggcg tggcctgggc gggactgggg agtggcgagc cctcagatcc
    10081 tgcatataag cagctgcttt ttgcctgtac tgggtctctc tggttagacc agatctgagc
    10141 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
    10201 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
    10261 acccttttag tcagtgtgga aaatctctag cagtagtagt tcatgtcatc ttattattca
    10321 gtatttataa cttgcaaaga aatgaatatc agagagtgag aggccttgac attgctagcg
    10381 tttaccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
    10441 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
    10501 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
    10561 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
    10621 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
    10681 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
    10741 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
    10801 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
    10861 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
    10921 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
    10981 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
    11041 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
    11101 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
    11161 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
    11221 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
    11281 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
    11341 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
    11401 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
    11461 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
    11521 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
    11581 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
    11641 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
    11701 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
    11761 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
    11821 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
    11881 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
    11941 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
    12001 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
    12061 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
    12121 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
    12181 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
    12241 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
    12301 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
    12361 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
    12421 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
    12481 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
    12541 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atcaacttgt
    12601 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
    12661 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
    12721 tctggatcaa ctggataact caagctaacc aaaatcatcc caaacttccc accccatacc
    12781 ctattaccac tgccaattac ctgtggtttc atttactcta aacctgtgat tcctctgaat
    12841 tattttcatt ttaaagaaat tgtatttgtt aaatatgtac tacaaactta gtagt
//
