LOCUS       sequence-344994-       11749 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inactivated Cas9 (dCas9) fusion to VPR for the purpose of targeted 
            activation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11749)
  TITLE     Brzezinski Dual Guide Donor/Activator/Repressor plasmids
REFERENCE   2  (bases 1 to 11749)
  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: pX458-Ef1a-dCas9-VPR-H2B-GFP (CRISPRa)
FEATURES             Location/Qualifiers
     source          1..11749
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        268..343
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        591..1773
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          822..1764
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1721..1741
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1779..1788
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1785..1811,1812..1814,1815..4298,4299..4304,4305..4370,
                     4371..4373,4374..5888)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=Cas9m4
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the mutations D10A, D839A, 
                     H840A, and N863A (Mali et al., 2013)"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVAAIVPQSFLKDDSIDNKVLTRSDKARGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5901..5921
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    5958..5982
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             6012..6161
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             6186..6206
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6636..6992
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALL"
     CDS             7011..7580
                     /codon_start=1
                     /product="transcriptional activation domain from the human 
                     herpesvirus 4 (Epstein-Barr virus) replication and 
                     transcription activator Rta/BRLF1 (Hardwick et al., 1992; 
                     Chavez et al., 2015)"
                     /label=Rta AD
                     /translation="RDSREGMFLPKPEAGSAISDVFEGREVCQPKRIRPFHPPGSPWAN
                     RPLPASLAPTPTGPVHEPVGSLTPAPVPQPLDPAPAVTPEASHLLEDPDEETSQAVKAL
                     REMADTVIPQKEEAAICGQMDLSHPPPRGHLDELTTTLESMTEDLNLDSPLTPELNEIL
                     DTFLNDECLLHAMHISTGLSIFDTSLF"
     CDS             7596..7649
                     /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             7650..8027
                     /codon_start=1
                     /gene="HIST1H2BB"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPSKSAPAPKKGSKKAITKAQKKDGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAVTKYTSSK"
     regulatory      8040..8049
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(8046..8048,8049..8051,8052..8762)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(8046..8048,8049..8051,8052..8762)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(8091..8112)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(8352..8371)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     8699..8720
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(8790..8807)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8796..9003
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   9012..9152
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   9012..9141
                     /label=AAV2 ITR
     rep_origin      9227..9682
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9314..9333)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9524..9545
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9699..9718)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     9818..9840
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(9878..9896)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9964..10068
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10069..10137,10138..10929)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(10287..10306)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11100..11688
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11589..11608
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ggtcttcgag aagacctgtt ttagagctag aaatagcaag ttaaaataag
      301 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg ttttagagct
      361 agaaatagca agttaaaata aggctagtcc gtttttagcg cgtgcgccaa ttctgcagac
      421 aaatggctct agaggtacca gtaattcata caaaaggact cgcccctgcc ttggggaatc
      481 ccagggaccg tcgttaaact cccactaacg tagaacccag agatcgctgc gttcccgccc
      541 cctcacccgc ccgctctcgt catcactgag gtggagaaga gcatgcgtga ggctccggtg
      601 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      661 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      721 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      781 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      841 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacgc
      901 ccctggctgc agtacgtgat tcttgatccc gagcttcggg ttggaagtgg gtgggagagt
      961 tcgaggcctt gcgcttaagg agccccttcg cctcgtgctt gagttgaggc ctggcttggg
     1021 cgctggggcc gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc tgctttcgat
     1081 aagtctctag ccatttaaaa tttttgatga cctgctgcga cgcttttttt ctggcaagat
     1141 agtcttgtaa atgcgggcca agatctgcac actggtattt cggtttttgg ggccgcgggc
     1201 ggcgacgggg cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg
     1261 ccaccgagaa tcggacgggg gtagtctcaa gctggccggc ctgctctggt gcctggcctc
     1321 gcgccgccgt gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc accagttgcg
     1381 tgagcggaaa gatggccgct tcccggccct gctgcaggga gctcaaaatg gaggacgcgg
     1441 cgctcgggag agcgggcggg tgagtcaccc acacaaagga aaagggcctt tccgtcctca
     1501 gccgtcgctt catgtgactc cacggagtac cgggcgccgt ccaggcacct cgattagttc
     1561 tcgagctttt ggagtacgtc gtctttaggt tggggggagg ggttttatgc gatggagttt
     1621 ccccacactg agtgggtgga gactgaagtt aggccagctt ggcacttgat gtaattctcc
     1681 ttggaatttg ccctttttga gtttggatct tggttcattc tcaagcctca gacagtggtt
     1741 caaagttttt ttcttccatt tcaggtgtcg tgaccggtgc caccatggac aagaagtact
     1801 ccattgggct cgctatcggc acaaacagcg tcggctgggc cgtcattacg gacgagtaca
     1861 aggtgccgag caaaaaattc aaagttctgg gcaataccga tcgccacagc ataaagaaga
     1921 acctcattgg cgccctcctg ttcgactccg gggagacggc cgaagccacg cggctcaaaa
     1981 gaacagcacg gcgcagatat acccgcagaa agaatcggat ctgctacctg caggagatct
     2041 ttagtaatga gatggctaag gtggatgact ctttcttcca taggctggag gagtcctttt
     2101 tggtggagga ggataaaaag cacgagcgcc acccaatctt tggcaatatc gtggacgagg
     2161 tggcgtacca tgaaaagtac ccaaccatat atcatctgag gaagaagctt gtagacagta
     2221 ctgataaggc tgacttgcgg ttgatctatc tcgcgctggc gcatatgatc aaatttcggg
     2281 gacacttcct catcgagggg gacctgaacc cagacaacag cgatgtcgac aaactcttta
     2341 tccaactggt tcagacttac aatcagcttt tcgaagagaa cccgatcaac gcatccggag
     2401 ttgacgccaa agcaatcctg agcgctaggc tgtccaaatc ccggcggctc gaaaacctca
     2461 tcgcacagct ccctggggag aagaagaacg gcctgtttgg taatcttatc gccctgtcac
     2521 tcgggctgac ccccaacttt aaatctaact tcgacctggc cgaagatgcc aagcttcaac
     2581 tgagcaaaga cacctacgat gatgatctcg acaatctgct ggcccagatc ggcgaccagt
     2641 acgcagacct ttttttggcg gcaaagaacc tgtcagacgc cattctgctg agtgatattc
     2701 tgcgagtgaa cacggagatc accaaagctc cgctgagcgc tagtatgatc aagcgctatg
     2761 atgagcacca ccaagacttg actttgctga aggcccttgt cagacagcaa ctgcctgaga
     2821 agtacaagga aattttcttc gatcagtcta aaaatggcta cgccggatac attgacggcg
     2881 gagcaagcca ggaggaattt tacaaattta ttaagcccat cttggaaaaa atggacggca
     2941 ccgaggagct gctggtaaag cttaacagag aagatctgtt gcgcaaacag cgcactttcg
     3001 acaatggaag catcccccac cagattcacc tgggcgaact gcacgctatc ctcaggcggc
     3061 aagaggattt ctaccccttt ttgaaagata acagggaaaa gattgagaaa atcctcacat
     3121 ttcggatacc ctactatgta ggccccctcg cccggggaaa ttccagattc gcgtggatga
     3181 ctcgcaaatc agaagagacc atcactccct ggaacttcga ggaagtcgtg gataaggggg
     3241 cctctgccca gtccttcatc gaaaggatga ctaactttga taaaaatctg cctaacgaaa
     3301 aggtgcttcc taaacactct ctgctgtacg agtacttcac agtttataac gagctcacca
     3361 aggtcaaata cgtcacagaa gggatgagaa agccagcatt cctgtctgga gagcagaaga
     3421 aagctatcgt ggacctcctc ttcaagacga accggaaagt taccgtgaaa cagctcaaag
     3481 aggactattt caaaaagatt gaatgtttcg actctgttga aatcagcgga gtggaggatc
     3541 gcttcaacgc atccctggga acgtatcacg atctcctgaa aatcattaaa gacaaggact
     3601 tcctggacaa tgaggagaac gaggacattc ttgaggacat tgtcctcacc cttacgttgt
     3661 ttgaagatag ggagatgatt gaagaacgct tgaaaactta cgctcatctc ttcgacgaca
     3721 aagtcatgaa acagctcaag aggcgccgat atacaggatg ggggcggctg tcaagaaaac
     3781 tgatcaatgg gatccgagac aagcagagtg gaaagacaat cctggatttt cttaagtccg
     3841 atggatttgc caaccggaac ttcatgcagt tgatccatga tgactctctc acctttaagg
     3901 aggacatcca gaaagcacaa gtttctggcc agggggacag tctgcacgag cacatcgcta
     3961 atcttgcagg tagcccagct atcaaaaagg gaatactgca gaccgttaag gtcgtggatg
     4021 aactcgtcaa agtaatggga aggcataagc ccgagaatat cgttatcgag atggcccgag
     4081 agaaccaaac tacccagaag ggacagaaga acagtaggga aaggatgaag aggattgaag
     4141 agggtataaa agaactgggg tcccaaatcc ttaaggaaca cccagttgaa aacacccagc
     4201 ttcagaatga gaagctctac ctgtactacc tgcagaacgg cagggacatg tacgtggatc
     4261 aggaactgga catcaatcgg ctctccgact acgacgtggc tgctatcgtg ccccagtctt
     4321 ttctcaaaga tgattctatt gataataaag tgttgacaag atccgataaa gctagaggga
     4381 agagtgataa cgtcccctca gaagaagttg tcaagaaaat gaaaaattat tggcggcagc
     4441 tgctgaacgc caaactgatc acacaacgga agttcgataa tctgactaag gctgaacgag
     4501 gtggcctgtc tgagttggat aaagccggct tcatcaaaag gcagcttgtt gagacacgcc
     4561 agatcaccaa gcacgtggcc caaattctcg attcacgcat gaacaccaag tacgatgaaa
     4621 atgacaaact gattcgagag gtgaaagtta ttactctgaa gtctaagctg gtctcagatt
     4681 tcagaaagga ctttcagttt tataaggtga gagagatcaa caattaccac catgcgcatg
     4741 atgcctacct gaatgcagtg gtaggcactg cacttatcaa aaaatatccc aagcttgaat
     4801 ctgaatttgt ttacggagac tataaagtgt acgatgttag gaaaatgatc gcaaagtctg
     4861 agcaggaaat aggcaaggcc accgctaagt acttctttta cagcaatatt atgaattttt
     4921 tcaagaccga gattacactg gccaatggag agattcggaa gcgaccactt atcgaaacaa
     4981 acggagaaac aggagaaatc gtgtgggaca agggtaggga tttcgcgaca gtccggaagg
     5041 tcctgtccat gccgcaggtg aacatcgtta aaaagaccga agtacagacc ggaggcttct
     5101 ccaaggaaag tatcctcccg aaaaggaaca gcgacaagct gatcgcacgc aaaaaagatt
     5161 gggaccccaa gaaatacggc ggattcgatt ctcctacagt cgcttacagt gtactggttg
     5221 tggccaaagt ggagaaaggg aagtctaaaa aactcaaaag cgtcaaggaa ctgctgggca
     5281 tcacaatcat ggagcgatca agcttcgaaa aaaaccccat cgactttctc gaggcgaaag
     5341 gatataaaga ggtcaaaaaa gacctcatca ttaagcttcc caagtactct ctctttgagc
     5401 ttgaaaacgg ccggaaacga atgctcgcta gtgcgggcga gctgcagaaa ggtaacgagc
     5461 tggcactgcc ctctaaatac gttaatttct tgtatctggc cagccactat gaaaagctca
     5521 aagggtctcc cgaagataat gagcagaagc agctgttcgt ggaacaacac aaacactacc
     5581 ttgatgagat catcgagcaa ataagcgagt tctccaaaag agtgatcctc gccgacgcta
     5641 acctcgataa ggtgctttct gcttacaata agcacaggga taagcccatc agggagcagg
     5701 cagaaaacat tatccacttg tttactctga ccaacttggg cgcgcctgca gccttcaagt
     5761 acttcgacac caccatagac agaaagcggt acacctctac aaaggaggtc ctggacgcca
     5821 cactgattca tcagtcaatt acggggctct atgaaacaag aatcgacctc tctcagctcg
     5881 gtggagacag cagggctgac cccaagaaga agaggaaggt gtcgccaggg atccgtcgac
     5941 ttgacgcgtt gatatcaaca agtttgtaca aaaaagcagg ctacaaagag gccagcggtt
     6001 ccggacgggc tgacgcattg gacgattttg atctggatat gctgggaagt gacgccctcg
     6061 atgattttga ccttgacatg cttggttcgg atgcccttga tgactttgac ctcgacatgc
     6121 tcggcagtga cgcccttgat gatttcgacc tggacatgct gattaactca agaagttccg
     6181 gatctccgaa aaagaaacgc aaagttggta gccagtacct gcccgacacc gacgaccggc
     6241 accggatcga ggaaaagcgg aagcggacct acgagacatt caagagcatc atgaagaagt
     6301 cccccttcag cggccccacc gaccctagac ctccacctag aagaatcgcc gtgcccagca
     6361 gatccagcgc cagcgtgcca aaacctgccc cccagcctta ccccttcacc agcagcctga
     6421 gcaccatcaa ctacgacgag ttccctacca tggtgttccc cagcggccag atctctcagg
     6481 cctctgctct ggctccagcc cctcctcagg tgctgcctca ggctcctgct cctgcaccag
     6541 ctccagccat ggtgtctgca ctggctcagg caccagcacc cgtgcctgtg ctggctcctg
     6601 gacctccaca ggctgtggct ccaccagccc ctaaacctac acaggccggc gagggcacac
     6661 tgtctgaagc tctgctgcag ctgcagttcg acgacgagga tctgggagcc ctgctgggaa
     6721 acagcaccga tcctgccgtg ttcaccgacc tggccagcgt ggacaacagc gagttccagc
     6781 agctgctgaa ccagggcatc cctgtggccc ctcacaccac cgagcccatg ctgatggaat
     6841 accccgaggc catcacccgg ctcgtgacag gcgctcagag gcctcctgat ccagctcctg
     6901 cccctctggg agcaccaggc ctgcctaatg gactgctgtc tggcgacgag gacttcagct
     6961 ctatcgccga tatggatttc tcagccttgc tgggctctgg cagcggcagc cgggattcca
     7021 gggaagggat gtttttgccg aagcctgagg ccggctccgc tattagtgac gtgtttgagg
     7081 gccgcgaggt gtgccagcca aaacgaatcc ggccatttca tcctccagga agtccatggg
     7141 ccaaccgccc actccccgcc agcctcgcac caacaccaac cggtccagta catgagccag
     7201 tcgggtcact gaccccggca ccagtccctc agccactgga tccagcgccc gcagtgactc
     7261 ccgaggccag tcacctgttg gaggatcccg atgaagagac gagccaggct gtcaaagccc
     7321 ttcgggagat ggccgatact gtgattcccc agaaggaaga ggctgcaatc tgtggccaaa
     7381 tggacctttc ccatccgccc ccaaggggcc atctggatga gctgacaacc acacttgagt
     7441 ccatgaccga ggatctgaac ctggactcac ccctgacccc ggaattgaac gagattctgg
     7501 ataccttcct gaacgacgag tgcctcttgc atgccatgca tatcagcaca ggactgtcca
     7561 tcttcgacac atctctgttt gaattcggca gtggagaggg cagaggaagt ctgctaacat
     7621 gcggtgacgt cgaggagaat cctggcccaa tgcctgaacc ctctaagtct gctccagccc
     7681 ctaaaaaggg ttctaagaag gctatcacta aggcgcagaa gaaggatggt aagaagcgta
     7741 agcgcagccg caaggagagc tattctatct atgtgtacaa ggttctgaag caggtccacc
     7801 ccgacaccgg catctcatcc aaggccatgg ggatcatgaa ctccttcgtc aacgacatct
     7861 tcgagcgcat cgcgggcgag gcttctcgcc tggctcacta caataagcgc tcgaccatca
     7921 cctccaggga gattcagacg gctgtgcgcc tgctgctgcc tggggagctg gctaagcatg
     7981 ctgtgtccga gggcactaag gcagttacca agtacactag ctctaaggat ccaccggtcg
     8041 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     8101 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     8161 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     8221 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     8281 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     8341 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     8401 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     8461 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     8521 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     8581 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     8641 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     8701 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     8761 aggaattcta actagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     8821 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     8881 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     8941 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gagaatagca ggcatgctgg
     9001 ggagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
     9061 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
     9121 agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac
     9181 gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc
     9241 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     9301 gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     9361 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     9421 ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag
     9481 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     9541 actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg
     9601 atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     9661 aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca
     9721 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     9781 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     9841 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     9901 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     9961 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
    10021 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
    10081 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
    10141 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
    10201 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
    10261 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
    10321 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
    10381 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
    10441 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
    10501 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
    10561 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
    10621 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
    10681 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
    10741 gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg cggtatcatt
    10801 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
    10861 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
    10921 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
    10981 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
    11041 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
    11101 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
    11161 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
    11221 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
    11281 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
    11341 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
    11401 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
    11461 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
    11521 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
    11581 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
    11641 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
    11701 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
