LOCUS       sequence-373287-       15219 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  dCas9 effector fusion for manipulating transcription.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15219)
  AUTHORS   Tycko J, Van MV, Aradhana, DelRosso N, Ye H, Yao D, Valbuena R, 
            Vaughan-Jackson A, Xu X, Ludwig C, Spees K, Liu K, Gu M, Khare V, 
            Mukund AX, Suzuki PH, Arana S, Zhang C, Du PP, Ornstein TS, Hess GT,
            Kamber RA, Qi LS, Khalil AS, Bintu L, Bassik MC
  TITLE     Development of compact transcriptional effectors using 
            high-throughput measurements in diverse contexts.
  JOURNAL   Nat Biotechnol. 2024 Nov 1. doi: 10.1038/s41587-024-02442-6.
  PUBMED    39487265
REFERENCE   2  (bases 1 to 15219)
  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: "Nat 
            Biotechnol. 2024 Nov 1. doi: 10.1038/s41587-024-02442-6."
COMMENT     Sequence Label: pJT258 Lenti
            pEF-dCas9-3XFLAG-N+F+Z-T2A-tagBFP...
FEATURES             Location/Qualifiers
     source          1..15219
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(55..74)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        246..625
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        627..825
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     780..800
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             843..1023
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1070..1195
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1688..1921
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2106..2150
                     /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    2448..2565
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2695..3873
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2926..3864
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3821..3841
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3889..3936
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(3937..3960,3961..3963,3964..6519,6520..6522,
                     6523..8037)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             8092..8112
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8128..8193
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8659..8712
                     /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             9439..9495
                     /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             9496..9894
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    9919..10507
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9972..9992)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(10390..10401)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(10510..10526)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10511..10527)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             11032..11212
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(11238..11255)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    11244..11468
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      11514..11942
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11601..11620)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11811..11832
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(11951..11971)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        11956..12285
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      12136..12271
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     12198..12217
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        12333..12380
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             12399..12773
                     /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    12903..13024
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12940..12959)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     12994..13013
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(13073..13089)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(13073..13089)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(13086..13108)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    13097..13113
                     /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(13121..13127,13128..13145,13146..13151))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    13166..13187
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(13304..13321)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(13475..14063)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(13555..13574)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(14234..15025,15026..15094))
                     /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     14857..14876
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(15095..15199)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
       61 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
      121 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
      181 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
      241 gcgttgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
      301 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
      361 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
      421 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
      481 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
      541 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
      601 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
      661 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      721 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      781 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagcgcgt tttgcctgta
      841 ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
      901 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
      961 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
     1021 gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg
     1081 caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac
     1141 gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat
     1201 taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa
     1261 aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa
     1321 tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc
     1381 ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg
     1441 tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga
     1501 gcaaaacaaa agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag
     1561 gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac
     1621 cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag
     1681 tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag
     1741 cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga
     1801 acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca
     1861 tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc
     1921 tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta
     1981 gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca
     2041 gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc
     2101 aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt
     2161 ttaacataac aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg
     2221 taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt
     2281 caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa
     2341 tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg aacggatcgg
     2401 cactgcgtgc gccaattctg cagacaaatg gcagtattca tccacaattt taaaagaaaa
     2461 ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc aacagacata
     2521 caaactaaag aattacaaaa acaaattaca aaaattcaaa attttcgggt ttattacagg
     2581 gacagcagag atccagtttg gttaattagc tagctgcaaa gatggataaa gttttaaaca
     2641 gagaggaatc tttgcagcta atggaccttc taggtcttga aaggagtggg aattggctcc
     2701 ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg
     2761 gtcggcaatt gaaccggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc
     2821 gtgtactggc tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc
     2881 gccgtgaacg ttctttttcg caacgggttt gccgccagaa cacaggtaag tgccgtgtgt
     2941 ggttcccgcg ggcctggcct ctttacgggt tatggccctt gcgtgccttg aattacttcc
     3001 acctggctgc agtacgtgat tcttgatccc gagcttcggg ttggaagtgg gtgggagagt
     3061 tcgaggcctt gcgcttaagg agccccttcg cctcgtgctt gagttgaggc ctggcctggg
     3121 cgctggggcc gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc tgctttcgat
     3181 aagtctctag ccatttaaaa tttttgatga cctgctgcga cgcttttttt ctggcaagat
     3241 agtcttgtaa atgcgggcca agatctgcac actggtattt cggtttttgg ggccgcgggc
     3301 ggcgacgggg cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg
     3361 ccaccgagaa tcggacgggg gtagtctcaa gctggccggc ctgctctggt gcctggcctc
     3421 gcgccgccgt gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc accagttgcg
     3481 tgagcggaaa gatggccgct tcccggccct gctgcaggga gctcaaaatg gaggacgcgg
     3541 cgctcgggag agcgggcggg tgagtcaccc acacaaagga aaagggcctt tccgtcctca
     3601 gccgtcgctt catgtgactc cacggagtac cgggcgccgt ccaggcacct cgattagttc
     3661 tcgagctttt ggagtacgtc gtctttaggt tggggggagg ggttttatgc gatggagttt
     3721 ccccacactg agtgggtgga gactgaagtt aggccagctt ggcacttgat gtaattctcc
     3781 ttggaatttg ccctttttga gtttggatct tggttcattc tcaagcctca gacagtggtt
     3841 caaagttttt ttcttccatt tcaggtgtcg tgacgtacgg ccaccatgaa aaggccggcg
     3901 gccacgaaaa aggccggcca ggcaaaaaag aaaaaggaca agaagtacag catcggcctg
     3961 gccatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc
     4021 aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga
     4081 gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga
     4141 agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag
     4201 atggccaagg tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag
     4261 gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac
     4321 gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc
     4381 gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg
     4441 atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg
     4501 cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag
     4561 gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg
     4621 cccggcgaga agaagaatgg cctgttcggc aacctgattg ccctgagcct gggcctgacc
     4681 cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac
     4741 acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg
     4801 tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac
     4861 accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac
     4921 caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag
     4981 attttcttcg accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag
     5041 gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg
     5101 ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc
     5161 atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt
     5221 tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc
     5281 tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc
     5341 gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag
     5401 agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc
     5461 aagcacagcc tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac
     5521 gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg
     5581 gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc
     5641 aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc
     5701 tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat
     5761 gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga
     5821 gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag
     5881 cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc
     5941 atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc
     6001 aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag
     6061 aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc
     6121 agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa
     6181 gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc
     6241 acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa
     6301 gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag
     6361 aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac
     6421 atcaaccggc tgtccgacta cgatgtggac cacatcgtgc ctcagagctt tctgaaggac
     6481 gactccatcg acaacaaggt gctgaccaga agcgacaagg cccggggcaa gagcgacaac
     6541 gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc
     6601 aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc
     6661 gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag
     6721 cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg
     6781 atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat
     6841 ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg
     6901 aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg
     6961 tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc
     7021 ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag
     7081 attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc
     7141 ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg
     7201 ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct
     7261 atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag
     7321 aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg
     7381 gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg
     7441 gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa
     7501 gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc
     7561 cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc
     7621 tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc
     7681 gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc
     7741 atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa
     7801 gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc
     7861 atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc
     7921 accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac
     7981 cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacagc
     8041 gctggaggag gtggaagcgg aggaggagga agcggaggag gaggtagcgg acctaagaaa
     8101 aagaggaagg tggcggccgc tggatccgac tacaaagacc atgacggtga ttataaagat
     8161 catgacatcg actacaagga tgacgatgac aagggtggat ccggatcagg ctccgagggc
     8221 cagagcgacg agagagccct gctggaccag ctgcacaccc tgctgagcaa caccgacgcc
     8281 accggcctgg aggagatcga cagagccctg ggcatccccg agctggtgaa ccagggccag
     8341 gccctggagc ccaagcaggg aggaggaggt agcggacacg agaagttccc cagcgacctg
     8401 gacctggaca tgttcaacgg cagcctggag tgcgacatgg agagcatcat cagaagcgag
     8461 ctgatggacg ccgacggcct ggacttcaac ttcgacagcg gaggaggagg tagcggattc
     8521 gtgaccctga aggacgtggg catggacttc accctgggcg actgggagca gctgggcctg
     8581 gagcagggcg acaccttctg ggacaccgcc ctggacaact gccaggacct gttcctgctg
     8641 gacccccccg gatctggaga gggaagggga agcctcctaa cttgcggaga tgtcgaggag
     8701 aatcctggcc caatggttag taagggagag gagttgataa aagagaatat gcacatgaaa
     8761 ctgtatatgg aagggacagt ggacaaccat cacttcaagt gcacctctga gggagaagga
     8821 aagccatacg aaggcaccca gactatgcga attaaggtcg tggagggagg gcctctcccc
     8881 ttcgccttcg atattcttgc aacatctttc ctctacggga gcaaaacctt catcaatcac
     8941 acacaaggca tacctgactt ttttaagcag agttttccag agggctttac ctgggagcgt
     9001 gtgactacat acgaagatgg gggtgtgctg acggctacac aggatacttc acttcaggac
     9061 ggttgtctga tctacaacgt caagatcagg ggggtcaatt tcacctcgaa tggaccagta
     9121 atgcaaaaaa agaccctggg atgggaagct tttactgaaa ctctctatcc cgccgacggc
     9181 ggcctcgaag gcagaaacga tatggctttg aagctggttg gcgggtccca tctaattgcc
     9241 aatgccaaaa ccacatatag gagcaaaaag cctgcaaaga acctgaagat gccgggtgtg
     9301 tactatgttg actatagatt agagcggatc aaagaggcaa ataacgaaac gtacgtggag
     9361 cagcacgagg tagccgtggc gcgctactgc gatttaccct ccaaactggg ccataaactg
     9421 aactgtacag gaagcggagc tactaacttc agcctgctga agcaggctgg cgacgtggaa
     9481 gaaaacccag gacctatggc caagcctttg tctcaagaag aatccaccct cattgaaaga
     9541 gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc cagcgcagct
     9601 ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac tgggggacct
     9661 tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa cctgacttgt
     9721 atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg gtgccgacag
     9781 gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga tggacagccg
     9841 acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg ctaagaattc
     9901 gatatcaagc ttatcggtaa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     9961 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
    10021 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
    10081 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt
    10141 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
    10201 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
    10261 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg
    10321 tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
    10381 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
    10441 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
    10501 tccccgcatc gataccgtcg acctcgagac ctagaaaaac atggagcaat cacaagtagc
    10561 aatacagcag ctaccaatgc tgattgtgcc tggctagaag cacaagagga ggaggaggtg
    10621 ggttttccag tcacacctca ggtaccttta agaccaatga cttacaaggc agctgtagat
    10681 cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga
    10741 caagatatcc ttgatctgtg gatctaccac acacaaggct acttccctga ttggcagaac
    10801 tacacaccag ggccagggat cagatatcca ctgacctttg gatggtgcta caagctagta
    10861 ccagttgagc aagagaaggt agaagaagcc aatgaaggag agaacacccg cttgttacac
    10921 cctgtgagcc tgcatgggat ggatgacccg gagagagaag tattagagtg gaggtttgac
    10981 agccgcctag catttcatca catggcccga gagctgcatc cggactgtac tgggtctctc
    11041 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
    11101 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
    11161 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagggcccgt
    11221 ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
    11281 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
    11341 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
    11401 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
    11461 ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc
    11521 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
    11581 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
    11641 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
    11701 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
    11761 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
    11821 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
    11881 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
    11941 ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
    12001 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
    12061 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
    12121 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
    12181 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc
    12241 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct
    12301 tgtatatcca ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg gcatagtata
    12361 tcggcatagt ataatacgac aaggtgagga actaaaccat ggccaagttg accagtgccg
    12421 ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga gttctggacc gaccggctcg
    12481 ggttctcccg ggacttcgtg gaggacgact tcgccggtgt ggtccgggac gacgtgaccc
    12541 tgttcatcag cgcggtccag gaccaggtgg tgccggacaa caccctggcc tgggtgtggg
    12601 tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt cgtgtccacg aacttccggg
    12661 acgcctccgg gccggccatg accgagatcg gcgagcagcc gtgggggcgg gagttcgccc
    12721 tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc
    12781 tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
    12841 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
    12901 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
    12961 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
    13021 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
    13081 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
    13141 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
    13201 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
    13261 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
    13321 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
    13381 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
    13441 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
    13501 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
    13561 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
    13621 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
    13681 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
    13741 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
    13801 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
    13861 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
    13921 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
    13981 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
    14041 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
    14101 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
    14161 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
    14221 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
    14281 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
    14341 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
    14401 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
    14461 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
    14521 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
    14581 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
    14641 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
    14701 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
    14761 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
    14821 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
    14881 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
    14941 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
    15001 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
    15061 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
    15121 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
    15181 aacaaatagg ggttccgcgc acatttcccc gaaaagtgc
//
