LOCUS       sequence-8481-ex        9984 bp    DNA     circular SYN 15-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9984)
  AUTHORS   Deans TL, Cantor CR, Collins JJ
  TITLE     A tunable genetic switch based on RNAi and repressor proteins for 
            regulating gene expression in mammalian cells.
  JOURNAL   Cell. 2007 Jul 27. 130(2):363-72.
  PUBMED    17662949
REFERENCE   2  (bases 1 to 9984)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell. 2007 
            Jul 27. 130(2):363-72."
COMMENT     Sequence Label: LTRi with neomycin
FEATURES             Location/Qualifiers
     source          1..9984
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        462..765
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        766..969
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             1022..2101
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(1041..1060)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2117..2137
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          2299..2364
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     polyA_signal    2523..2644
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2677..2693)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2701..2717
                     /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(2725..2731,2732..2749,2750..2755))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2770..2791
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     3026..3042
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3049..3067
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3162..3180
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     primer_bind     complement(3192..3211)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(3362..3382)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     promoter        complement(3528..3546)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3529..3546)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3564..3580)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3588..3604
                     /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(3612..3618,3619..3636,3637..3642))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3657..3678
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3795..3812)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     4020..4036
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        4103..4406
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4407..4610
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    4728..4747
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     protein_bind    4771..4790
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     protein_bind    4814..4833
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     primer_bind     5277..5296
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     promoter        5324..5342
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5353..5973
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(5426..5445)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     polyA_signal    6015..6136
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(6177..6210)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     polyA_signal    6232..6695
                     /label=PGK poly(A) signal
                     /note="mouse phosphoglycerate kinase 1 polyadenylation 
                     signal"
     CDS             complement(6705..7508)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6816..6835)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     7426..7445
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(7522..8021)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(7570..7589)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     7963..7986
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     protein_bind    complement(8037..8070)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     rep_origin      complement(8165..8753)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8245..8264)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8924..9715,9716..9784))
                     /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     9547..9566
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9785..9889)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     9957..9975
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt ctagttatta atagtaatca
      421 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
      481 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
      541 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
      601 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
      661 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
      721 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
      781 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
      841 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
      901 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      961 agcagagctg gtttagtgaa ccgtcagatc ggatcctcta gagtcgacca ttgccgccac
     1021 catgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
     1081 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
     1141 ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca
     1201 gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt
     1261 cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga
     1321 acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag
     1381 tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg
     1441 cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat
     1501 tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca
     1561 gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg
     1621 ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg
     1681 gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac
     1741 tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc
     1801 cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc
     1861 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
     1921 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
     1981 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
     2041 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     2101 gagcagcctg aggcctccta agaagaagag gaaggtttga gcgcaacgca attaatgtaa
     2161 gttagctcac tcattaggca ccccaggctt tacactttat gcttccgacc tgcagcccct
     2221 tggatctttg tgaaggaacc ttacttctgt ggtgtgacat aattggacaa actacctaca
     2281 gagatttaaa gctctaaggt aaatataaaa tttttaagtg tataatgtgt taaactactg
     2341 attctaattg tttgtgtatt ttagatcaca gtcccaaggc tcatttcagg cccctcagtc
     2401 ctcacagtct gttcatgatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
     2461 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     2521 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     2581 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     2641 cttatcatgt ctgggcccaa gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     2701 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     2761 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     2821 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     2881 tttgcgtatt gggcgctctt ccgctgcaac tgttgggaag ggcgatcggt gcgggcctct
     2941 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg
     3001 ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaattgta atacgactca
     3061 ctatagggcg aattgggcct ttttctacac aaatcagcga ttttctcttg aaaaatcgct
     3121 gatttgtgta gcggtgtttc gtcctttcca caagatatat aactctatca atgatagagt
     3181 actttcaagt tacggtaagc atatgatagt ccattttaaa acataatttt aaaactgcaa
     3241 actacccaag aaattattac tttctacgtc acgtattttg tactaatatc tttgtgttta
     3301 cagtcaaatt aattctaatt atctctctaa cagccttgta tcgtatatgc aaatatgaag
     3361 gaatcatggg aaataggccc tcttcctgcc cgaccttggc gcgcgctcgg cgcgcggtca
     3421 cgctccgtca cgtggtgcgt tttgcctaat cactagtgaa ttcgcggccg cctgcaggtc
     3481 gaccatatgg gagagctccc aacgcgttgg atgcatagct tgagtattct atagtgtcac
     3541 ctaaatagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     3601 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     3661 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     3721 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     3781 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     3841 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     3901 aagaacatgg ccggcttaat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
     3961 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
     4021 taaaacgacg gccagtgaat tctagttatt aatagtaatc aattacgggg tcattagttc
     4081 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
     4141 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
     4201 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
     4261 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
     4321 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
     4381 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
     4441 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
     4501 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
     4561 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga
     4621 accgtcagat cggatcctct agagtcgacc tgcaggatct aagcttggac aaactaccta
     4681 cagagattta aagctctaag gtaaatataa aatttactag gttgtggaat tgtgagcgct
     4741 cacaattcca cagtcgaccc taggttgtgg aattgtgagc gctcacaatt ccacagtcga
     4801 ccctaggttg tggaattgtg agcgctcaca attccacagt cgaccctagt gtataatgtg
     4861 ttaaactacg gatccgtctc ccattaggcc tacaatggtg agacaagtag ccaacaggga
     4921 agggttgcaa atatcatttg ggcacaccta tgataatatt gatgaagcag acagtattca
     4981 gcaagtaact gagaggtggg aagctcaaag ccaaagtcct aatgtgcagt caggtgaatt
     5041 tattgaaaaa tttgaggctc ctggtggtgc aaatcaaaga actgctcctc agggatccta
     5101 attgtttgtg tattttagat tccaaccaag cttgcggccg ctcaggagct aaggaagcta
     5161 aaatggagaa aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag
     5221 aacgggcccc tctgctaacc atgttcatgc cttcttcttt ttcctacagc tcctgggcaa
     5281 cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttgtaatacg actcactata
     5341 gggcgaattg atatgtctag attagataaa agtaaagtga ttaacagcgc attagagctg
     5401 cttaatgagg tcggaatcga aggtttaaca acccgtaaac tcgcccagaa gctaggtgta
     5461 gagcagccta cattgtattg gcatgtaaaa aataagcggg ctttgctcga cgccttagcc
     5521 attgagatgt tagataggca ccatactcac ttttgccctt tagaagggga aagctggcaa
     5581 gattttttac gtaataacgc taaaagtttt agatgtgctt tactaagtca tcgcgatgga
     5641 gcaaaagtac atttaggtac acggcctaca gaaaaacagt atgaaactct cgaaaatcaa
     5701 ttagcctttt tatgccaaca aggtttttca ctagagaatg cattatatgc actcagcgct
     5761 gtggggcatt ttactttagg ttgcgtattg gaagatcaag agcatcaagt cgctaaagaa
     5821 gaaagggaaa cacctactac tgatagtatg ccgccattat tacgacaagc tatcgaatta
     5881 tttgatcacc aaggtgcaga gccagccttc ttattcggcc ttgaattgat catatgcgga
     5941 ttagaaaaac aacttaaatg tgaaagtggg tccgcgtaca gcggatcccg ggaattcaga
     6001 tcttattaaa gcagaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     6061 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     6121 tcatcaatgt atcttatcat gtctggtcga atccatcaca ctggcggccg ctcgacgaag
     6181 ttcctatact ttctagagaa taggaacttc cctcgacggg tatcgataag cttctgatgg
     6241 aattagaact tggcaaaaca atactgnaga atgaagtgta tgttggaaca gaggctgctg
     6301 atctcgttct tcaggctatg aaactgacac atttggnaan cacagtactt agaaccacaa
     6361 agtgggaatc aagagaaaaa caatgatccc acgagagatc tatagatcta tagatcatga
     6421 gtgggaggaa tgagctggcc cttaatttgg ttttgcttgt ttaaattatg atatccaact
     6481 atgaaacatt atcataaagc aatagtaaag agccttcagt aaagagcagg catttatcta
     6541 atcccacccc acccccaccc ccgtagctcc aatccttcca ttcaaaatgt aggtactctg
     6601 ttctcaccct tcttaacaaa gtatgacagg aaaaacttcc attttagtgg acatctttat
     6661 tgtttaatag atcatcaatt tctgcagact tacagcggat cccctcagaa gaactcgtca
     6721 agaaggcgat agaaggcgat gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg
     6781 aagcggtcag cccattcgcc gccaagctct tcagcaatat cacgggtagc caacgctatg
     6841 tcctgatagc ggtccgccac acccagccgg ccacagtcga tgaatccaga aaagcggcca
     6901 ttttccacca tgatattcgg caagcaggca tcgccatggg tcacgacgag atcctcgccg
     6961 tcgggcatgc gcgccttgag cctggcgaac agttcggctg gcgcgagccc ctgatgctct
     7021 tcgtccagat catcctgatc gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg
     7081 cgatgtttcg cttggtggtc gaatgggcag gtagccggat caagcgtatg cagccgccgc
     7141 attgcatcag ccatgatgga tactttctcg gcaggagcaa ggtgagatga caggagatcc
     7201 tgccccggca cttcgcccaa tagcagccag tcccttcccg cttcagtgac aacgtcgagc
     7261 acagctgcgc aaggaacgcc cgtcgtggcc agccacgata gccgcgctgc ctcgtcctgc
     7321 agttcattca gggcaccgga caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct
     7381 gacagccgga acacggcggc atcagagcag ccgattgtct gttgtgccca gtcatagccg
     7441 aatagcctct ccacccaagc ggccggagaa cctgcgtgca atccatcttg ttcaatggcc
     7501 gatcccatat tggctgcagg tcgaaaggcc cggagatgag gaagaggaga acagcgcggc
     7561 agacgtgcgc ttttgaagcg tgcagaatgc cgggcctccg gaggaccttc gggcgcccgc
     7621 cccgcccctg agcccgcccc tgagcccgcc cccggaccca ccccttccca gcctctgagc
     7681 ccagaaagcg aaggagcaaa gctgctattg gccgctgccc caaaggccta cccgcttcca
     7741 ttgctcagcg gtgctgtcca tctgcacgag actagtgaga cgtgctactt ccatttgtca
     7801 cgtcctgcac gacgcgagct gcggggcggg ggggaacttc ctgactaggg gaggagtaga
     7861 aggtggcgcg aaggggccac caaagaacgg agccggttgg cgcctaccgg tggatgtgga
     7921 atgtgtgcga ggccagaggc cacttgtgta gcgccaagtg cccagcgggg ctgctaaagc
     7981 gcatgctcca gactgccttg ggaaaagcgc ctcccctacc cggtagaatt ggccgcgaag
     8041 ttcctatact ttctagagaa taggaacttc ggccaaaacg tgtcgagtct agagggccac
     8101 atgtcatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     8161 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     8221 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     8281 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     8341 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     8401 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     8461 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     8521 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     8581 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     8641 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     8701 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     8761 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     8821 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     8881 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     8941 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     9001 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     9061 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     9121 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     9181 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     9241 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     9301 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     9361 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     9421 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     9481 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     9541 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     9601 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     9661 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     9721 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     9781 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     9841 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     9901 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     9961 ggcgtatcac gaggcccttt cgtc
//
