LOCUS       sequence-387166-       15264 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 15264)
  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 15264)
  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: pJT341 Lenti pEF-dCas9-3XFLAG-KRAB[WSR7EEE] + ...
FEATURES             Location/Qualifiers
     source          1..15264
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(3..107)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(182..201)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        373..752
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        754..952
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     907..927
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             970..1150
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1197..1322
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1815..2048
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2233..2277
                     /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    2575..2692
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2822..4000
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3053..3991
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3948..3968
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4016..4063
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(4064..4087,4088..4090,4091..6646,6647..6649,
                     6650..8164)
                     /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             8219..8239
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8255..8320
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8372..8554
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="TLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGYQ
                     LTKPDVILRLEKGEEP"
     CDS             8831..8884
                     /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             9611..9667
                     /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             9668..10066
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    10091..10679
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(10144..10164)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(10562..10573)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(10682..10698)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10683..10699)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             11204..11384
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(11410..11427)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    11416..11640
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      11686..12114
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11773..11792)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11983..12004
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(12123..12143)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        12128..12457
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      12308..12443
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     12370..12389
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        12505..12552
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             12571..12945
                     /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    13075..13196
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(13112..13131)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     13166..13185
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(13245..13261)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(13245..13261)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(13258..13280)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    13269..13285
                     /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(13293..13299,13300..13317,13318..13323))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    13338..13359
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(13476..13493)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(13647..14235)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(13727..13746)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(14406..15197,15198..15264,1..2))
                     /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     15029..15048
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
       61 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
      121 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
      181 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
      241 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
      301 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
      361 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
      421 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      481 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      541 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      601 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      661 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      721 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
      781 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
      841 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
      901 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agcgcgtttt
      961 gcctgtactg ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta
     1021 gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc
     1081 cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa
     1141 atctctagca gtggcgcccg aacagggact tgaaagcgaa agggaaacca gaggagctct
     1201 ctcgacgcag gactcggctt gctgaagcgc gcacggcaag aggcgagggg cggcgactgg
     1261 tgagtacgcc aaaaattttg actagcggag gctagaagga gagagatggg tgcgagagcg
     1321 tcagtattaa gcgggggaga attagatcgc gatgggaaaa aattcggtta aggccagggg
     1381 gaaagaaaaa atataaatta aaacatatag tatgggcaag cagggagcta gaacgattcg
     1441 cagttaatcc tggcctgtta gaaacatcag aaggctgtag acaaatactg ggacagctac
     1501 aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca gtagcaaccc
     1561 tctattgtgt gcatcaaagg atagagataa aagacaccaa ggaagcttta gacaagatag
     1621 aggaagagca aaacaaaagt aagaccaccg cacagcaagc ggccgctgat cttcagacct
     1681 ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa
     1741 attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa
     1801 agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg
     1861 ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag
     1921 cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc
     1981 tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa
     2041 cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg
     2101 aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag
     2161 tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa
     2221 aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg
     2281 aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga
     2341 ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag
     2401 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
     2461 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
     2521 ggatcggcac tgcgtgcgcc aattctgcag acaaatggca gtattcatcc acaattttaa
     2581 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
     2641 agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta
     2701 ttacagggac agcagagatc cagtttggtt aattagctag ctgcaaagat ggataaagtt
     2761 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
     2821 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
     2881 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
     2941 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
     3001 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
     3061 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
     3121 tacttccacc tggctgcagt acgtgattct tgatcccgag cttcgggttg gaagtgggtg
     3181 ggagagttcg aggccttgcg cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg
     3241 gcctgggcgc tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc
     3301 tttcgataag tctctagcca tttaaaattt ttgatgacct gctgcgacgc tttttttctg
     3361 gcaagatagt cttgtaaatg cgggccaaga tctgcacact ggtatttcgg tttttggggc
     3421 cgcgggcggc gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg
     3481 agcgcggcca ccgagaatcg gacgggggta gtctcaagct ggccggcctg ctctggtgcc
     3541 tggcctcgcg ccgccgtgta tcgccccgcc ctgggcggca aggctggccc ggtcggcacc
     3601 agttgcgtga gcggaaagat ggccgcttcc cggccctgct gcagggagct caaaatggag
     3661 gacgcggcgc tcgggagagc gggcgggtga gtcacccaca caaaggaaaa gggcctttcc
     3721 gtcctcagcc gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca ggcacctcga
     3781 ttagttctcg agcttttgga gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat
     3841 ggagtttccc cacactgagt gggtggagac tgaagttagg ccagcttggc acttgatgta
     3901 attctccttg gaatttgccc tttttgagtt tggatcttgg ttcattctca agcctcagac
     3961 agtggttcaa agtttttttc ttccatttca ggtgtcgtga cgtacggcca ccatgaaaag
     4021 gccggcggcc acgaaaaagg ccggccaggc aaaaaagaaa aaggacaaga agtacagcat
     4081 cggcctggcc atcggcacca actctgtggg ctgggccgtg atcaccgacg agtacaaggt
     4141 gcccagcaag aaattcaagg tgctgggcaa caccgaccgg cacagcatca agaagaacct
     4201 gatcggagcc ctgctgttcg acagcggcga aacagccgag gccacccggc tgaagagaac
     4261 cgccagaaga agatacacca gacggaagaa ccggatctgc tatctgcaag agatcttcag
     4321 caacgagatg gccaaggtgg acgacagctt cttccacaga ctggaagagt ccttcctggt
     4381 ggaagaggat aagaagcacg agcggcaccc catcttcggc aacatcgtgg acgaggtggc
     4441 ctaccacgag aagtacccca ccatctacca cctgagaaag aaactggtgg acagcaccga
     4501 caaggccgac ctgcggctga tctatctggc cctggcccac atgatcaagt tccggggcca
     4561 cttcctgatc gagggcgacc tgaaccccga caacagcgac gtggacaagc tgttcatcca
     4621 gctggtgcag acctacaacc agctgttcga ggaaaacccc atcaacgcca gcggcgtgga
     4681 cgccaaggcc atcctgtctg ccagactgag caagagcaga cggctggaaa atctgatcgc
     4741 ccagctgccc ggcgagaaga agaatggcct gttcggcaac ctgattgccc tgagcctggg
     4801 cctgaccccc aacttcaaga gcaacttcga cctggccgag gatgccaaac tgcagctgag
     4861 caaggacacc tacgacgacg acctggacaa cctgctggcc cagatcggcg accagtacgc
     4921 cgacctgttt ctggccgcca agaacctgtc cgacgccatc ctgctgagcg acatcctgag
     4981 agtgaacacc gagatcacca aggcccccct gagcgcctct atgatcaaga gatacgacga
     5041 gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta
     5101 caaagagatt ttcttcgacc agagcaagaa cggctacgcc ggctacattg acggcggagc
     5161 cagccaggaa gagttctaca agttcatcaa gcccatcctg gaaaagatgg acggcaccga
     5221 ggaactgctc gtgaagctga acagagagga cctgctgcgg aagcagcgga ccttcgacaa
     5281 cggcagcatc ccccaccaga tccacctggg agagctgcac gccattctgc ggcggcagga
     5341 agatttttac ccattcctga aggacaaccg ggaaaagatc gagaagatcc tgaccttccg
     5401 catcccctac tacgtgggcc ctctggccag gggaaacagc agattcgcct ggatgaccag
     5461 aaagagcgag gaaaccatca ccccctggaa cttcgaggaa gtggtggaca agggcgcttc
     5521 cgcccagagc ttcatcgagc ggatgaccaa cttcgataag aacctgccca acgagaaggt
     5581 gctgcccaag cacagcctgc tgtacgagta cttcaccgtg tataacgagc tgaccaaagt
     5641 gaaatacgtg accgagggaa tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc
     5701 catcgtggac ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga
     5761 ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt
     5821 caacgcctcc ctgggcacat accacgatct gctgaaaatt atcaaggaca aggacttcct
     5881 ggacaatgag gaaaacgagg acattctgga agatatcgtg ctgaccctga cactgtttga
     5941 ggacagagag atgatcgagg aacggctgaa aacctatgcc cacctgttcg acgacaaagt
     6001 gatgaagcag ctgaagcggc ggagatacac cggctggggc aggctgagcc ggaagctgat
     6061 caacggcatc cgggacaagc agtccggcaa gacaatcctg gatttcctga agtccgacgg
     6121 cttcgccaac agaaacttca tgcagctgat ccacgacgac agcctgacct ttaaagagga
     6181 catccagaaa gcccaggtgt ccggccaggg cgatagcctg cacgagcaca ttgccaatct
     6241 ggccggcagc cccgccatta agaagggcat cctgcagaca gtgaaggtgg tggacgagct
     6301 cgtgaaagtg atgggccggc acaagcccga gaacatcgtg atcgaaatgg ccagagagaa
     6361 ccagaccacc cagaagggac agaagaacag ccgcgagaga atgaagcgga tcgaagaggg
     6421 catcaaagag ctgggcagcc agatcctgaa agaacacccc gtggaaaaca cccagctgca
     6481 gaacgagaag ctgtacctgt actacctgca gaatgggcgg gatatgtacg tggaccagga
     6541 actggacatc aaccggctgt ccgactacga tgtggaccac atcgtgcctc agagctttct
     6601 gaaggacgac tccatcgaca acaaggtgct gaccagaagc gacaaggccc ggggcaagag
     6661 cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag aactactggc ggcagctgct
     6721 gaacgccaag ctgattaccc agagaaagtt cgacaatctg accaaggccg agagaggcgg
     6781 cctgagcgaa ctggataagg ccggcttcat caagagacag ctggtggaaa cccggcagat
     6841 cacaaagcac gtggcacaga tcctggactc ccggatgaac actaagtacg acgagaatga
     6901 caagctgatc cgggaagtga aagtgatcac cctgaagtcc aagctggtgt ccgatttccg
     6961 gaaggatttc cagttttaca aagtgcgcga gatcaacaac taccaccacg cccacgacgc
     7021 ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc tggaaagcga
     7081 gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag atgatcgcca agagcgagca
     7141 ggaaatcggc aaggctaccg ccaagtactt cttctacagc aacatcatga actttttcaa
     7201 gaccgagatt accctggcca acggcgagat ccggaagcgg cctctgatcg agacaaacgg
     7261 cgaaaccggg gagatcgtgt gggataaggg ccgggatttt gccaccgtgc ggaaagtgct
     7321 gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa
     7381 agagtctatc ctgcccaaga ggaacagcga taagctgatc gccagaaaga aggactggga
     7441 ccctaagaag tacggcggct tcgacagccc caccgtggcc tattctgtgc tggtggtggc
     7501 caaagtggaa aagggcaagt ccaagaaact gaagagtgtg aaagagctgc tggggatcac
     7561 catcatggaa agaagcagct tcgagaagaa tcccatcgac tttctggaag ccaagggcta
     7621 caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag tactccctgt tcgagctgga
     7681 aaacggccgg aagagaatgc tggcctctgc cggcgaactg cagaagggaa acgaactggc
     7741 cctgccctcc aaatatgtga acttcctgta cctggccagc cactatgaga agctgaaggg
     7801 ctcccccgag gataatgagc agaaacagct gtttgtggaa cagcacaagc actacctgga
     7861 cgagatcatc gagcagatca gcgagttctc caagagagtg atcctggccg acgctaatct
     7921 ggacaaagtg ctgtccgcct acaacaagca ccgggataag cccatcagag agcaggccga
     7981 gaatatcatc cacctgttta ccctgaccaa tctgggagcc cctgccgcct tcaagtactt
     8041 tgacaccacc atcgaccgga agaggtacac cagcaccaaa gaggtgctgg acgccaccct
     8101 gatccaccag agcatcaccg gcctgtacga gacacggatc gacctgtctc agctgggagg
     8161 cgacagcgct ggaggaggtg gaagcggagg aggaggaagc ggaggaggag gtagcggacc
     8221 taagaaaaag aggaaggtgg cggccgctgg atccgactac aaagaccatg acggtgatta
     8281 taaagatcat gacatcgact acaaggatga cgatgacaag ggtggatccg gatcaggctc
     8341 cgatgccaag tccttgaccg ctgaggagga gaccctggtc actttcaagg acgtttttgt
     8401 ggatttcacg agggaagaat ggaaactact agacacagca caacaaatag tatacagaaa
     8461 cgtaatgcta gaaaactaca aaaacctagt aagcctagga taccaactaa caaaaccaga
     8521 cgtaatacta agactagaaa aaggagaaga accaggagga ggaggtagcg gacctaggtt
     8581 gaagaatgtt gatagatcca ccgctcaaca gttggccgtg actgtgggaa atgtcacagt
     8641 cattatcaca gattttaagg aaaaaaccgg aggaggaggt agcggaatca aggaggagcc
     8701 cctggacgac tacgacggca gctcatcttc ccccgaaaat gatgatctct ttatgatgcc
     8761 acggattgta aatgttacat ccctcgctac ggaaggcgga ttggtcgata tgggtggttc
     8821 tggatctgga gagggaaggg gaagcctcct aacttgcgga gatgtcgagg agaatcctgg
     8881 cccaatggtt agtaagggag aggagttgat aaaagagaat atgcacatga aactgtatat
     8941 ggaagggaca gtggacaacc atcacttcaa gtgcacctct gagggagaag gaaagccata
     9001 cgaaggcacc cagactatgc gaattaaggt cgtggaggga gggcctctcc ccttcgcctt
     9061 cgatattctt gcaacatctt tcctctacgg gagcaaaacc ttcatcaatc acacacaagg
     9121 catacctgac ttttttaagc agagttttcc agagggcttt acctgggagc gtgtgactac
     9181 atacgaagat gggggtgtgc tgacggctac acaggatact tcacttcagg acggttgtct
     9241 gatctacaac gtcaagatca ggggggtcaa tttcacctcg aatggaccag taatgcaaaa
     9301 aaagaccctg ggatgggaag cttttactga aactctctat cccgccgacg gcggcctcga
     9361 aggcagaaac gatatggctt tgaagctggt tggcgggtcc catctaattg ccaatgccaa
     9421 aaccacatat aggagcaaaa agcctgcaaa gaacctgaag atgccgggtg tgtactatgt
     9481 tgactataga ttagagcgga tcaaagaggc aaataacgaa acgtacgtgg agcagcacga
     9541 ggtagccgtg gcgcgctact gcgatttacc ctccaaactg ggccataaac tgaactgtac
     9601 aggaagcgga gctactaact tcagcctgct gaagcaggct ggcgacgtgg aagaaaaccc
     9661 aggacctatg gccaagcctt tgtctcaaga agaatccacc ctcattgaaa gagcaacggc
     9721 tacaatcaac agcatcccca tctctgaaga ctacagcgtc gccagcgcag ctctctctag
     9781 cgacggccgc atcttcactg gtgtcaatgt atatcatttt actgggggac cttgtgcaga
     9841 actcgtggtg ctgggcactg ctgctgctgc ggcagctggc aacctgactt gtatcgtcgc
     9901 gatcggaaat gagaacaggg gcatcttgag cccctgcgga cggtgccgac aggtgcttct
     9961 cgatctgcat cctgggatca aagccatagt gaaggacagt gatggacagc cgacggcagt
    10021 tgggattcgt gaattgctgc cctctggtta tgtgtgggag ggctaagaat tcgatatcaa
    10081 gcttatcggt aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
    10141 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
    10201 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
    10261 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
    10321 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
    10381 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
    10441 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc
    10501 ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
    10561 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
    10621 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca
    10681 tcgataccgt cgacctcgag acctagaaaa acatggagca atcacaagta gcaatacagc
    10741 agctaccaat gctgattgtg cctggctaga agcacaagag gaggaggagg tgggttttcc
    10801 agtcacacct caggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca
    10861 ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatat
    10921 ccttgatctg tggatctacc acacacaagg ctacttccct gattggcaga actacacacc
    10981 agggccaggg atcagatatc cactgacctt tggatggtgc tacaagctag taccagttga
    11041 gcaagagaag gtagaagaag ccaatgaagg agagaacacc cgcttgttac accctgtgag
    11101 cctgcatggg atggatgacc cggagagaga agtattagag tggaggtttg acagccgcct
    11161 agcatttcat cacatggccc gagagctgca tccggactgt actgggtctc tctggttaga
    11221 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
    11281 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
    11341 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagggccc gtttaaaccc
    11401 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
    11461 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
    11521 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
    11581 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
    11641 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
    11701 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
    11761 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
    11821 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
    11881 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
    11941 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
    12001 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
    12061 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct
    12121 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
    12181 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
    12241 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
    12301 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
    12361 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta
    12421 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc
    12481 cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata
    12541 gtataatacg acaaggtgag gaactaaacc atggccaagt tgaccagtgc cgttccggtg
    12601 ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc
    12661 cgggacttcg tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc
    12721 agcgcggtcc aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc
    12781 ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc
    12841 gggccggcca tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac
    12901 ccggccggca actgcgtgca cttcgtggcc gaggagcagg actgacacgt gctacgagat
    12961 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
    13021 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
    13081 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
    13141 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
    13201 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
    13261 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
    13321 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
    13381 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
    13441 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
    13501 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
    13561 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
    13621 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
    13681 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
    13741 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
    13801 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
    13861 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
    13921 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
    13981 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
    14041 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
    14101 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
    14161 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
    14221 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
    14281 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
    14341 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
    14401 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
    14461 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
    14521 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
    14581 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
    14641 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
    14701 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
    14761 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
    14821 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
    14881 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
    14941 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
    15001 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
    15061 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
    15121 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
    15181 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
    15241 gcgacacgga aatgttgaat actc
//
