LOCUS       sequence-236365-       14418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Cas9, 2xNES and 2 tandem ERT2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14418)
  AUTHORS   Zhao C, Zhao Y, Zhang J, Lu J, Chen L, Zhang Y, Ying Y, Xu J, Wei S,
            Wang Y
  TITLE     HIT-Cas9: A CRISPR/Cas9 Genome-Editing Device under Tight and 
            Effective Drug Control.
  JOURNAL   Mol Ther Nucleic Acids. 2018 Dec 7;13:208-219. doi: 
            10.1016/j.omtn.2018.08.022. Epub 2018 Sep 1.
  PUBMED    30312845
REFERENCE   2  (bases 1 to 14418)
  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: "Mol Ther 
            Nucleic Acids."; date: "2018-12-7"; volume: "13:208-219. doi"; 
            pages: " 10.1016/j.omtn.2018.08.022. Epub 2018 Sep 1"
COMMENT     Sequence Label: EF1a-Cas9-2XNES-2ERT2
FEATURES             Location/Qualifiers
     source          1..14418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        130..341
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      1205..1214
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1226..5326
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMTKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5465..6400
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
                     LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     CDS             6452..7387
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
                     LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     CDS             6500..7393
                     /codon_start=1
                     /gene="ESR1"
                     /product="estrogen nuclear receptor alpha ligand-binding 
                     domain"
                     /label=ER-LBD
                     /translation="KRSKKNSLALSLTADQMVSALLDAEPPILYSEYDPTRPFSEASMM
                     GLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEILMIGLVWRSMEHPVKL
                     LFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEFVCLKSIILLNSGVYTF
                     LSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRLAQLLLILSHIRHMSNK
                     GMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEETDQSHLATAGSTSSHSL
                     QKYYITGEAEGFPATA"
     primer_bind     complement(7537..7557)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7542..7871
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7722..7857
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             7881..8255
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     misc_feature    8341..8929
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8394..8414)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8812..8823)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9016..9249
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    9321..9442
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      9461..9596
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(9616..9635)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(9617..9635)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9645..9662)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(9645..9661)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9654..9676)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9803..10258
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9890..9909)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10100..10121
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        10284..10388
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10389..10457,10458..11249)
                     /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     complement(10607..10626)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11420..12008
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11909..11928
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     12162..12179
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    12296..12317
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(12332..12337,12338..12355,12356..12362)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12370..12386
                     /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)."
     primer_bind     12375..12397
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     12394..12410
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     12394..12410
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     12429..12449
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        12431..12449
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        12477..12703
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             12704..12884
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    12931..13056
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    13549..13782
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             13967..14011
                     /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    14278..14395
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 atagcccggg ctcgaccata tgataagctt tgcaaagatg gataaagttt taaacagaga
       61 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      121 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      181 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      241 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      301 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      361 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      421 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      481 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      541 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      601 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      661 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      721 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      781 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      841 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
      901 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
      961 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1021 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1081 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1141 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctcctttc
     1201 tagagccacc atgggtggcg cgcctgacaa gaagtacagc atcggcctgg acatcggcac
     1261 caactctgtg ggctgggccg tgatcaccga cgagtacaag gtgcccagca agaaattcaa
     1321 ggtgctgggc aacaccgacc ggcacagcat caagaagaac ctgatcggag ccctgctgtt
     1381 cgacagcggc gaaacagccg aggccacccg gctgaagaga accgccagaa gaagatacac
     1441 cagacggaag aaccggatct gctatctgca agagatcttc agcaacgaga tgaccaaggt
     1501 ggacgacagc ttcttccaca gactggaaga gtccttcctg gtggaagagg ataagaagca
     1561 cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc
     1621 caccatctac cacctgagaa agaaactggt ggacagcacc gacaaggccg acctgcggct
     1681 gatctatctg gccctggccc acatgatcaa gttccggggc cacttcctga tcgagggcga
     1741 cctgaacccc gacaacagcg acgtggacaa gctgttcatc cagctggtgc agacctacaa
     1801 ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc
     1861 tgccagactg agcaagagca gacggctgga aaatctgatc gcccagctgc ccggcgagaa
     1921 gaagaatggc ctgttcggaa acctgattgc cctgagcctg ggcctgaccc ccaacttcaa
     1981 gagcaacttc gacctggccg aggatgccaa actgcagctg agcaaggaca cctacgacga
     2041 cgacctggac aacctgctgg cccagatcgg cgaccagtac gccgacctgt ttctggccgc
     2101 caagaacctg tccgacgcca tcctgctgag cgacatcctg agagtgaaca ccgagatcac
     2161 caaggccccc ctgagcgcct ctatgatcaa gagatacgac gagcaccacc aggacctgac
     2221 cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag tacaaagaga ttttcttcga
     2281 ccagagcaag aacggctacg ccggctacat tgacggcgga gccagccagg aagagttcta
     2341 caagttcatc aagcccatcc tggaaaagat ggacggcacc gaggaactgc tcgtgaagct
     2401 gaacagagag gacctgctgc ggaagcagcg gaccttcgac aacggcagca tcccccacca
     2461 gatccacctg ggagagctgc acgccattct gcggcggcag gaagattttt acccattcct
     2521 gaaggacaac cgggaaaaga tcgagaagat cctgaccttc cgcatcccct actacgtggg
     2581 ccctctggcc aggggaaaca gcagattcgc ctggatgacc agaaagagcg aggaaaccat
     2641 caccccctgg aacttcgagg aagtggtgga caagggcgct tccgcccaga gcttcatcga
     2701 gcggatgacc aacttcgata agaacctgcc caacgagaag gtgctgccca agcacagcct
     2761 gctgtacgag tacttcaccg tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg
     2821 aatgagaaag cccgccttcc tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt
     2881 caagaccaac cggaaagtga ccgtgaagca gctgaaagag gactacttca agaaaatcga
     2941 gtgcttcgac tccgtggaaa tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac
     3001 ataccacgat ctgctgaaaa ttatcaagga caaggacttc ctggacaatg aggaaaacga
     3061 ggacattctg gaagatatcg tgctgaccct gacactgttt gaggacagag agatgatcga
     3121 ggaacggctg aaaacctatg cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg
     3181 gcggagatac accggctggg gcaggctgag ccggaagctg atcaacggca tccgggacaa
     3241 gcagtccggc aagacaatcc tggatttcct gaagtccgac ggcttcgcca acagaaactt
     3301 catgcagctg atccacgacg acagcctgac ctttaaagag gacatccaga aagcccaggt
     3361 gtccggccag ggcgatagcc tgcacgagca cattgccaat ctggccggca gccccgccat
     3421 taagaagggc atcctgcaga cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg
     3481 gcacaagccc gagaacatcg tgatcgaaat ggccagagag aaccagacca cccagaaggg
     3541 acagaagaac agccgcgaga gaatgaagcg gatcgaagag ggcatcaaag agctgggcag
     3601 ccagatcctg aaagaacacc ccgtggaaaa cacccagctg cagaacgaga agctgtacct
     3661 gtactacctg cagaatgggc gggatatgta cgtggaccag gaactggaca tcaaccggct
     3721 gtccgactac gatgtggacc atatcgtgcc tcagagcttt ctgaaggacg actccatcga
     3781 caacaaggtg ctgaccagaa gcgacaagaa ccggggcaag agcgacaacg tgccctccga
     3841 agaggtcgtg aagaagatga agaactactg gcggcagctg ctgaacgcca agctgattac
     3901 ccagagaaag ttcgacaatc tgaccaaggc cgagagaggc ggcctgagcg aactggataa
     3961 ggccggcttc atcaagagac agctggtgga aacccggcag atcacaaagc acgtggcaca
     4021 gatcctggac tcccggatga acactaagta cgacgagaat gacaagctga tccgggaagt
     4081 gaaagtgatc accctgaagt ccaagctggt gtccgatttc cggaaggatt tccagtttta
     4141 caaagtgcgc gagatcaaca actaccacca cgcccacgac gcctacctga acgccgtcgt
     4201 gggaaccgcc ctgatcaaaa agtaccctaa gctggaaagc gagttcgtgt acggcgacta
     4261 caaggtgtac gacgtgcgga agatgatcgc caagagcgag caggaaatcg gcaaggctac
     4321 cgccaagtac ttcttctaca gcaacatcat gaactttttc aagaccgaga ttaccctggc
     4381 caacggcgag atccggaagc ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt
     4441 gtgggataag ggccgggatt ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa
     4501 tatcgtgaaa aagaccgagg tgcagacagg cggcttcagc aaagagtcta tcctgcccaa
     4561 gaggaacagc gataagctga tcgccagaaa gaaggactgg gaccctaaga agtacggcgg
     4621 cttcgacagc cccaccgtgg cctattctgt gctggtggtg gccaaagtgg aaaagggcaa
     4681 gtccaagaaa ctgaagagtg tgaaagagct gctggggatc accatcatgg aaagaagcag
     4741 cttcgagaag aatcccatcg actttctgga agccaagggc tacaaagaag tgaaaaagga
     4801 cctgatcatc aagctgccta agtactccct gttcgagctg gaaaacggcc ggaagagaat
     4861 gctggcctct gccggcgaac tgcagaaggg aaacgaactg gccctgccct ccaaatatgt
     4921 gaacttcctg tacctggcca gccactatga gaagctgaag ggctcccccg aggataatga
     4981 gcagaaacag ctgtttgtgg aacagcacaa gcactacctg gacgagatca tcgagcagat
     5041 cagcgagttc tccaagagag tgatcctggc cgacgctaat ctggacaaag tgctgtccgc
     5101 ctacaacaag caccgggata agcccatcag agagcaggcc gagaatatca tccacctgtt
     5161 taccctgacc aatctgggag cccctgccgc cttcaagtac tttgacacca ccatcgaccg
     5221 gaagaggtac accagcacca aagaggtgct ggacgccacc ctgatccacc agagcatcac
     5281 cggcctgtac gagacacgga tcgacctgtc tcagctggga ggcgacggag gtagtggagg
     5341 tactagttta gccttgaaat tagcaggtct tgatatcgct agcggtacct tagccttgaa
     5401 attagcaggt cttgatatca cgcgtgctag ccgcctgctg gaagatggcg atctcgagcc
     5461 atctgctgga gacatgagag ctgccaacct ttggccaagc ccgctcatga tcaaacgctc
     5521 taagaagaac agcctggcct tgtccctgac ggccgaccag atggtcagtg ccttgttgga
     5581 tgctgagccc cccatactct attccgagta tgatcctacc agacccttca gtgaagcttc
     5641 gatgatgggc ttactgacca acctggcaga cagggagctg gttcacatga tcaactgggc
     5701 gaagagggtg ccaggctttg tggatttgac cctccatgat caggtccacc ttctagaatg
     5761 tgcctggcta gagatcctga tgattggtct cgtctggcgc tccatggagc acccagtgaa
     5821 gctactgttt gctcctaact tgctcttgga caggaaccag ggaaaatgtg tagagggcat
     5881 ggtggagatc ttcgacatgc tgctggctac atcatctcgg ttccgcatga tgaatctgca
     5941 gggagaggag tttgtgtgcc tcaaatctat tattttgctt aattctggag tgtacacatt
     6001 tctgtccagc accctgaagt ctctggaaga gaaggaccat atccaccgag tcctggacaa
     6061 gatcacagac actttgatcc acctgatggc caaggcaggc ctgaccctgc agcagcagca
     6121 ccagcggctg gcccagctcc tcctcatcct ctcccacatc aggcacatga gtaacaaagg
     6181 catggagcat ctgtacagca tgaagtgcaa gaacgtggtg cccctctatg acctgctgct
     6241 ggaggcggcg gacgcccacc gcctacatgc gcccactagc cgtggagggg catccgtgga
     6301 ggagacggac caaagccact tggccactgc gggctctact tcatcgcatt ccttgcaaaa
     6361 gtattacatc acgggggagg cagagggttt ccctgccaca gctggatccc gcactagtcg
     6421 cctgctggaa gatggcgatc tcgagccatc tgctggagac atgagagctg ccaacctttg
     6481 gccaagcccg ctcatgatca aacgctctaa gaagaacagc ctggccttgt ccctgacggc
     6541 cgaccagatg gtcagtgcct tgttggatgc tgagcccccc atactctatt ccgagtatga
     6601 tcctaccaga cccttcagtg aagcttcgat gatgggctta ctgaccaacc tggcagacag
     6661 ggagctggtt cacatgatca actgggcgaa gagggtgcca ggctttgtgg atttgaccct
     6721 ccatgatcag gtccaccttc tagaatgtgc ctggctagag atcctgatga ttggtctcgt
     6781 ctggcgctcc atggagcacc cagtgaagct actgtttgct cctaacttgc tcttggacag
     6841 gaaccaggga aaatgtgtag agggcatggt ggagatcttc gacatgctgc tggctacatc
     6901 atctcggttc cgcatgatga atctgcaggg agaggagttt gtgtgcctca aatctattat
     6961 tttgcttaat tctggagtgt acacatttct gtccagcacc ctgaagtctc tggaagagaa
     7021 ggaccatatc caccgagtcc tggacaagat cacagacact ttgatccacc tgatggccaa
     7081 ggcaggcctg accctgcagc agcagcacca gcggctggcc cagctcctcc tcatcctctc
     7141 ccacatcagg cacatgagta acaaaggcat ggagcatctg tacagcatga agtgcaagaa
     7201 cgtggtgccc ctctatgacc tgctgctgga ggcggcggac gcccaccgcc tacatgcgcc
     7261 cactagccgt ggaggggcat ccgtggagga gacggaccaa agccacttgg ccactgcggg
     7321 ctctacttca tcgcattcct tgcaaaagta ttacatcacg ggggaggcag agggtttccc
     7381 tgccacagct tgattcgaag tcgacttgtt tattgcagct tataatggtt acaaataaag
     7441 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     7501 gtccaaactc atcaatgtat cttatgtcga ccctgtggaa tgtgtgtcag ttagggtgtg
     7561 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     7621 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     7681 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     7741 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     7801 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     7861 gcttttgcaa aaagcttacc atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc
     7921 gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc cgggacttcg
     7981 tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc agcgcggtcc
     8041 aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc
     8101 tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca
     8161 tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac ccggccggca
     8221 actgcgtgca cttcgtggcc gaggagcagg actgaccgat cgataaaata aaagatttta
     8281 tttagtctcc agaaaaaggg gggaatgaaa gaccccacct gtaggtttgg caagctcgac
     8341 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     8401 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     8461 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     8521 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     8581 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     8641 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     8701 ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc atggctgctc
     8761 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     8821 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     8881 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc tggaattcga
     8941 gctcagtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     9001 aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat ctgctttttg
     9061 cttgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     9121 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     9181 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     9241 tctctagcag tagtagttca tgtcatctta ttattcagta tttataactt gcaaagaaat
     9301 gaatatcaga gagtgagagg aacttgttta ttgcagctta taatggttac aaataaagca
     9361 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     9421 ccaaactcat caatgtatct tatcatgtct ggctctagct atcccgcccc taactccgcc
     9481 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     9541 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     9601 gacgtaccca attcgcccta tagtgagtcg tattacgcgc gctcactggc cgtcgtttta
     9661 caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc
     9721 cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     9781 cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     9841 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     9901 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     9961 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
    10021 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
    10081 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
    10141 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
    10201 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag
    10261 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
    10321 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
    10381 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
    10441 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
    10501 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
    10561 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
    10621 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
    10681 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
    10741 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
    10801 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
    10861 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
    10921 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
    10981 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
    11041 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
    11101 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
    11161 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
    11221 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
    11281 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
    11341 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
    11401 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
    11461 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
    11521 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
    11581 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
    11641 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
    11701 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
    11761 ccagcttgga gcgaacgacc tacaccaaac tgagatacct acagcgtgag ctatgagaaa
    11821 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
    11881 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
    11941 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
    12001 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
    12061 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
    12121 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
    12181 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
    12241 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
    12301 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
    12361 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
    12421 ccaagcgcgc aattaaccct cactaaaggg aacaaaagct ggagctgcaa gcttaatgta
    12481 gtcttatgca atactcttgt agtcttgcaa catggtaacg atgagttagc aacatgcctt
    12541 acaaggagag aaaaagcacc gtgcatgccg attggtggaa gtaaggtggt acgatcgtgc
    12601 cttattagga aggcaacaga cgggtctgac atggattgga cgaaccactg aattgccgca
    12661 ttgcagagat attgtattta agtgcctagc tcgatacata aacgggtctc tctggttaga
    12721 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
    12781 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
    12841 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg
    12901 gacttgaaag cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa
    12961 gcgcgcacgg caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc
    13021 ggaggctaga aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga
    13081 tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat
    13141 atagtatggg caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca
    13201 tcagaaggct gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa
    13261 gaacttagat cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag
    13321 ataaaagaca ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc
    13381 accgcacagc aagcggccgc tgatcttcag acctggagga ggagatatga gggacaattg
    13441 gagaagtgaa ttatataaat ataaagtagt aaaaattgaa ccattaggag tagcacccac
    13501 caaggcaaag agaagagtgg tgcagagaga aaaaagagca gtgggaatag gagctttgtt
    13561 ccttgggttc ttgggagcag caggaagcac tatgggcgca gcgtcaatga cgctgacggt
    13621 acaggccaga caattattgt ctggtatagt gcagcagcag aacaatttgc tgagggctat
    13681 tgaggcgcaa cagcatctgt tgcaactcac agtctggggc atcaagcagc tccaggcaag
    13741 aatcctggct gtggaaagat acctaaagga tcaacagctc ctggggattt ggggttgctc
    13801 tggaaaactc atttgcacca ctgctgtgcc ttggaatgct agttggagta ataaatctct
    13861 ggaacagatt tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac
    13921 aagcttaata cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga
    13981 attattggaa ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct
    14041 gtggtatata aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt
    14101 tgctgtactt tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac
    14161 ccacctccca accccgaggg gacccgacag gcccgaagga atagaagaag aaggtggaga
    14221 gagagacaga gacagatcca ttcgattagt gaacggatct cgacggtatc ggttaacttt
    14281 taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc
    14341 aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa attttatcga
    14401 tcacgagact agcctcga
//
