LOCUS       sequence-246444-        9819 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SP-Cas9 protein in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9819)
  AUTHORS   D'Astolfo DS, Pagliero RJ, Pras A, Karthaus WR, Clevers H, Prasad V,
            Lebbink RJ, Rehmann H, Geijsen N
  TITLE     Efficient Intracellular Delivery of Native Proteins.
  JOURNAL   Cell. 2015 Apr 23;161(3):674-690. doi: 10.1016/j.cell.2015.03.028.
  PUBMED    25910214
REFERENCE   2  (bases 1 to 9819)
  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"; doi: 
            "10.1016/j.cell.2015.03"; journalName: "Cell"; date: "2015-04-23- 
            23"; volume: "161"; issue: "3"; pages: "674-690"
COMMENT     Sequence Label: SP-Cas9
FEATURES             Location/Qualifiers
     source          1..9819
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(18..36)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        104..208
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(209..277,278..1069)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(427..446)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1240..1828
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1729..1748
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1982..1999
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2014..2153
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2079..2098)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2239..2261
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2255..2446)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3758..4840)
                     /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     4802..4821
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4841..4918)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5124..5143
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5227..5246
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5227..5245
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5246..5270
                     /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)."
     RBS             5285..5307
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5314..9417
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5314..9417
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             9442..9462
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             9475..9492
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(9546..9564)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      9560..9607
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(9782..9787,9788..9804,9805..9810))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
       61 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
      121 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
      181 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
      241 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
      301 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
      361 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
      421 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
      481 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
      541 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
      601 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
      661 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
      721 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
      781 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
      841 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
      901 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
      961 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     1021 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     1081 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     1141 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     1201 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     1261 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     1321 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     1381 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     1441 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     1501 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     1561 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     1621 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     1681 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     1741 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     1801 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     1861 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     1921 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     1981 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     2041 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     2101 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     2161 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     2221 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     2281 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     2341 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     2401 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     2461 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     2521 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     2581 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     2641 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     2701 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     2761 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     2821 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     2881 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc
     2941 gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt
     3001 gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt
     3061 tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg
     3121 caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca
     3181 tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag
     3241 gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct
     3301 ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat
     3361 ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc
     3421 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     3481 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     3541 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     3601 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     3661 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     3721 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     3781 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     3841 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     3901 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     3961 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     4021 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     4081 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     4141 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     4201 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     4261 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     4321 gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg
     4381 tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa
     4441 tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa
     4501 gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca
     4561 cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt
     4621 gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt
     4681 gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg
     4741 ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac
     4801 cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac
     4861 tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg
     4921 ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg
     4981 aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt
     5041 cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag
     5101 tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct
     5161 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg
     5221 cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa
     5281 taattttgtt taactttaag aaggagatat accatggata aaaaatacag cattggtctg
     5341 gacattggca cgaatagcgt tggttgggca gtgattaccg atgaatacaa agtcccgtcg
     5401 aaaaaattca aagtgctggg taacaccgat cgccatagca ttaagaaaaa cctgatcggt
     5461 gcgctgctgt ttgattctgg cgaaaccgcg gaagcaacgc gtctgaaacg taccgcacgt
     5521 cgccgttaca cgcgccgtaa aaatcgtatt tgctatctgc aggaaatctt tagcaacgaa
     5581 atggcgaaag tcgatgactc atttttccac cgcctggaag aatcgtttct ggtggaagaa
     5641 gataaaaaac atgaacgtca cccgattttc ggcaatatcg ttgatgaagt cgcgtaccat
     5701 gaaaaatatc cgacgattta ccacctgcgt aaaaaactgg tggattctac cgacaaagcc
     5761 gatctgcgcc tgatttatct ggcactggct catatgatca aatttcgtgg tcacttcctg
     5821 attgaaggcg acctgaaccc ggataatagt gacgtcgata aactgtttat tcagctggtg
     5881 caaacctata atcagctgtt cgaagaaaac ccgatcaatg caagtggtgt tgatgcgaaa
     5941 gccattctgt ccgctcgcct gagtaaatcc cgccgtctgg aaaacctgat tgcacagctg
     6001 ccgggtgaaa agaaaaacgg tctgtttggc aatctgatcg ctctgtcact gggcctgacg
     6061 ccgaacttta aatcgaattt cgacctggca gaagatgcta aactgcagct gagcaaagat
     6121 acctacgatg acgatctgga caacctgctg gcgcaaattg gcgaccagta tgccgacctg
     6181 tttctggcgg ccaaaaatct gtcagatgcc attctgctgt cggacatcct gcgcgtgaac
     6241 accgaaatca cgaaagcgcc gctgtcagcc tcgatgatta aacgctacga tgaacatcac
     6301 caggacctga ccctgctgaa agcactggtt cgtcagcaac tgccggaaaa atacaaagaa
     6361 attttctttg accaaagtaa aaatggttat gcaggctaca tcgatggcgg tgcttcccag
     6421 gaagaattct acaaattcat caaaccgatc ctggaaaaaa tggatggtac ggaagaactg
     6481 ctggtgaaac tgaatcgtga agatctgctg cgtaaacaac gcacctttga caacggtagc
     6541 attccgcatc agatccacct gggcgaactg catgcgattc tgcgccgtca ggaagatttt
     6601 tatccgttcc tgaaagacaa ccgtgaaaaa atcgaaaaaa tcctgacgtt tcgcatcccg
     6661 tattacgttg gtccgctggc acgtggtaat agccgcttcg catggatgac ccgcaaatct
     6721 gaagaaacca ttacgccgtg gaactttgaa gaagtggttg ataaaggcgc aagcgctcag
     6781 tcttttatcg aacgtatgac caatttcgat aaaaacctgc cgaatgaaaa agtgctgccg
     6841 aaacattctc tgctgtatga atactttacc gtttacaacg aactgacgaa agtgaaatat
     6901 gttaccgagg gtatgcgcaa accggcgttt ctgagtggcg aacagaaaaa agccattgtg
     6961 gatctgctgt tcaaaaccaa tcgtaaagtt acggtcaaac agctgaaaga agattacttc
     7021 aagaaaattg aatgtttcga cagcgtggaa atttctggtg ttgaagatcg tttcaacgcc
     7081 tctctgggca cctatcatga cctgctgaaa atcatcaaag acaaagattt tctggataac
     7141 gaagaaaacg aagacattct ggaagatatc gtgctgaccc tgacgctgtt cgaagatcgt
     7201 gaaatgattg aagaacgcct gaaaacgtac gcacacctgt ttgacgataa agttatgaaa
     7261 cagctgaaac gccgtcgcta taccggttgg ggccgtctga gccgcaaact gattaatggt
     7321 atccgcgata aacaatcagg caaaacgatt ctggatttcc tgaaatcgga cggctttgcc
     7381 aaccgtaatt tcatgcagct gatccatgac gattccctga cctttaaaga agacattcag
     7441 aaagcacaag tgtcaggtca aggcgattcg ctgcatgaac acattgcgaa cctggccggt
     7501 tcaccggcta tcaaaaaagg catcctgcag accgtgaaag tcgtggatga actggtgaaa
     7561 gttatgggtc gtcacaaacc ggaaaacatt gttatcgaaa tggcgcgcga aaatcagacc
     7621 acgcaaaaag gccagaaaaa ctcgcgtgaa cgcatgaaac gcattgaaga aggtatcaaa
     7681 gaactgggca gccagattct gaaagaacat ccggtcgaaa acacccagct gcaaaatgaa
     7741 aaactgtacc tgtattacct gcaaaatggt cgtgacatgt atgtggatca ggaactggac
     7801 atcaaccgcc tgtctgacta tgatgtcgac cacattgtgc cgcagagctt tctgaaagac
     7861 gattctatcg ataacaaagt tctgacccgt agtgataaaa accgcggcaa aagcgacaat
     7921 gtcccgtctg aagaagttgt gaagaaaatg aaaaactact ggcgtcaact gctgaatgcg
     7981 aaactgatta cgcagcgtaa attcgataac ctgaccaaag cggaacgcgg cggtctgtcc
     8041 gaactggata aagccggttt tatcaaacgt caactggttg aaacccgcca gattacgaaa
     8101 catgtcgccc agatcctgga ttcacgcatg aacacgaaat acgacgaaaa cgataaactg
     8161 atccgtgaag tcaaagtgat caccctgaaa agtaaactgg tttccgattt ccgtaaagac
     8221 tttcagttct acaaagtccg cgaaattaac aattaccatc acgcacacga tgcttatctg
     8281 aatgcagtgg ttggtaccgc tctgatcaaa aaatatccga aactggaaag cgaatttgtg
     8341 tatggcgatt acaaagtcta tgacgtgcgc aaaatgattg cgaaatccga acaggaaatc
     8401 ggcaaagcga ccgccaaata ctttttctat tcaaacatca tgaacttttt caaaaccgaa
     8461 attacgctgg caaatggtga aattcgtaaa cgcccgctga tcgaaaccaa cggtgaaacg
     8521 ggcgaaattg tgtgggataa aggccgtgac ttcgcgaccg ttcgcaaagt cctgtcgatg
     8581 ccgcaagtga atatcgtgaa gaaaaccgaa gtgcagacgg gcggttttag taaagaatcc
     8641 atcctgccga aacgtaacag cgataaactg attgcgcgca aaaaagattg ggacccgaaa
     8701 aaatacggcg gttttgatag tccgacggtt gcatattccg tcctggtcgt ggctaaagtc
     8761 gaaaaaggta aaagtaaaaa actgaaatcc gtgaaagaac tgctgggcat taccatcatg
     8821 gaacgtagct cttttgagaa aaacccgatt gacttcctgg aagccaaagg ttacaaagaa
     8881 gtgaaaaaag atctgatcat caaactgccg aaatatagcc tgttcgaact ggaaaacggc
     8941 cgtaaacgca tgctggcatc tgctggtgaa ctgcagaaag gcaatgaact ggcactgccg
     9001 agtaaatatg ttaactttct gtacctggct agccattatg aaaaactgaa aggttctccg
     9061 gaagataacg aacagaaaca actgttcgtc gaacaacata aacactacct ggatgaaatc
     9121 atcgaacaga tctcagaatt ctcgaaacgc gtgattctgg cggatgccaa tctggacaaa
     9181 gttctgagcg cgtataacaa acatcgtgat aaaccgattc gcgaacaggc cgaaaatatt
     9241 atccacctgt ttaccctgac gaacctgggc gcaccggcag cttttaaata cttcgatacc
     9301 acgatcgacc gtaaacgcta tacctcaacg aaagaagttc tggatgctac cctgattcat
     9361 caatcgatca ccggtctgta tgaaacgcgt attgatctga gtcagctggg cggtgacggc
     9421 ggtggctccg gcacccgtct gccgaaaaag aaacgcaaag tgggcggtgg ctcgcatcat
     9481 catcatcatc actaaggatc cggctgctaa caaagcccga aaggaagctg agttggctgc
     9541 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
     9601 ttttttgctg aaaggaggaa ctatatccgg atatcccgca agaggcccgg cagtaccggc
     9661 ataaccaagc ctatgcctac agcatccagg gtgacggtgc cgaggatgac gatgagcgca
     9721 ttgttagatt tcatacacgg tgcctgactg cgttagcaat ttaactgtga taaactaccg
     9781 cattaaagct tatcgatgat aagctgtcaa acatgagaa
//
