LOCUS       sequence-275015-       13136 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SIX1 knockout.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13136)
  TITLE     Kingsbury Lab Plasmids
REFERENCE   2  (bases 1 to 13136)
  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"
COMMENT     Sequence Label: pTJK643
FEATURES             Location/Qualifiers
     source          1..13136
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1443..1676
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1861..1905
                     /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    2203..2320
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2371..2619
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2371..2391
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2542..2561
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2641..2716
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2778..2989
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3008..3017
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3014..7117
                     /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             3014..7117
                     /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             7118..7165
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7166..7189
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7199..7255
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(7256..7258,7259..7261,7262..7972)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(7562..7581)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7909..7930
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    7991..8579
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8044..8064)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8462..8473)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8651..8884
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8910..8927)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8916..9140
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9186..9614
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9273..9292)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9483..9504
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9623..9643)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9628..9957
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9808..9943
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9870..9889
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10005..10052
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10071..10445
                     /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"
     polyA_signal    10575..10696
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10612..10631)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10666..10685
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10745..10761)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10745..10761)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10758..10780)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10769..10785
                     /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(10793..10799,10800..10817,10818..10823))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10838..10859
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10976..10993)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11147..11735)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11227..11246)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11906..12697,12698..12766))
                     /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     12529..12548
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12767..12871)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(12946..12965)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat
     1321 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta
     1381 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga
     1441 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca
     1501 atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat
     1561 ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag
     1621 cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg
     1681 atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg
     1741 agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa
     1801 attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa
     1861 aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac
     1921 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt
     1981 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca
     2041 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa
     2101 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg
     2161 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     2221 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2281 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     2341 cagagatcca gtttggttaa ttaaggtacc gagggcctat ttcccatgat tccttcatat
     2401 ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac tgtaaacaca
     2461 aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt
     2521 ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat
     2581 ttcttggctt tatatatctt gtggaaagga cgaaacaccg tgcttcaagg agaagtcgag
     2641 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
     2701 ggcaccgagt cggtgctttt ttgaattcgc tagctaggtc ttgaaaggag tgggaattgg
     2761 ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
     2821 aggggtcggc aattgatccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
     2881 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
     2941 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg accggttcta
     3001 gagcgctgcc accatggaca agaagtacag catcggcctg gacatcggca ccaactctgt
     3061 gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg
     3121 caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg
     3181 cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa
     3241 gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag
     3301 cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca
     3361 ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta
     3421 ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct
     3481 ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc
     3541 cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt
     3601 cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact
     3661 gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg
     3721 cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt
     3781 cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga
     3841 caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct
     3901 gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc
     3961 cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa
     4021 agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa
     4081 gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat
     4141 caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga
     4201 ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct
     4261 gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa
     4321 ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc
     4381 caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg
     4441 gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac
     4501 caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga
     4561 gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa
     4621 gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa
     4681 ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcttcga
     4741 ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca cataccacga
     4801 tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg aggacattct
     4861 ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg aggaacggct
     4921 gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc ggcggagata
     4981 caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca agcagtccgg
     5041 caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact tcatgcagct
     5101 gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg tgtccggcca
     5161 gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca ttaagaaggg
     5221 catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc
     5281 cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg gacagaagaa
     5341 cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca gccagatcct
     5401 gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc tgtactacct
     5461 gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc tgtccgacta
     5521 cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg acaacaaggt
     5581 gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg aagaggtcgt
     5641 gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta cccagagaaa
     5701 gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata aggccggctt
     5761 catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac agatcctgga
     5821 ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag tgaaagtgat
     5881 caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt acaaagtgcg
     5941 cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg tgggaaccgc
     6001 cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact acaaggtgta
     6061 cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta
     6121 cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg ccaacggcga
     6181 gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa
     6241 gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa
     6301 aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca agaggaacag
     6361 cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg gcttcgacag
     6421 ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca agtccaagaa
     6481 actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca gcttcgagaa
     6541 gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat
     6601 caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa tgctggcctc
     6661 tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg tgaacttcct
     6721 gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg agcagaaaca
     6781 gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga tcagcgagtt
     6841 ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa
     6901 gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt ttaccctgac
     6961 caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc ggaagaggta
     7021 caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca ccggcctgta
     7081 cgagacacgg atcgacctgt ctcagctggg aggcgacaag cgacctgccg ccacaaagaa
     7141 ggctggacag gctaagaaga agaaagatta caaagacgat gacgataagg gatccggcgc
     7201 aacaaacttc tctctgctga aacaagccgg agatgtcgaa gagaatcctg gaccgatggt
     7261 gtctaagggc gaagagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     7321 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     7381 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     7441 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     7501 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     7561 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     7621 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     7681 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     7741 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     7801 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     7861 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     7921 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaaacgcg
     7981 ttaagtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     8041 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     8101 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     8161 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     8221 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     8281 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     8341 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc
     8401 ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     8461 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     8521 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcg
     8581 tcgactttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa
     8641 aaggggggac tggaagggct aattcactcc caacgaagac aagatctgct ttttgcttgt
     8701 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     8761 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     8821 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     8881 agcagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     8941 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     9001 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     9061 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     9121 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
     9181 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     9241 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     9301 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     9361 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     9421 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     9481 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     9541 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     9601 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     9661 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     9721 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     9781 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     9841 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     9901 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     9961 ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac aattaatcat
    10021 cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc atggccaagt
    10081 tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga
    10141 ccgaccggct cgggttctcc cgggacttcg tggaggacga cttcgccggt gtggtccggg
    10201 acgacgtgac cctgttcatc agcgcggtcc aggaccaggt ggtgccggac aacaccctgg
    10261 cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca
    10321 cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag ccgtgggggc
    10381 gggagttcgc cctgcgcgac ccggccggca actgcgtgca cttcgtggcc gaggagcagg
    10441 actgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
    10501 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
    10561 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
    10621 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
    10681 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
    10741 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
    10801 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
    10861 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
    10921 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
    10981 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
    11041 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
    11101 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
    11161 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
    11221 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
    11281 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
    11341 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
    11401 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
    11461 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
    11521 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
    11581 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
    11641 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
    11701 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
    11761 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
    11821 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
    11881 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
    11941 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
    12001 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
    12061 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
    12121 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
    12181 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
    12241 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
    12301 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
    12361 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
    12421 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
    12481 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
    12541 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
    12601 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
    12661 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
    12721 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
    12781 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
    12841 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
    12901 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
    12961 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
    13021 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
    13081 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgtt
//
