LOCUS       sequence-495592-       12911 bp    DNA     circular SYN 17-DEC-2025
DEFINITION  Control nickase Cas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12911)
  AUTHORS   Sokirniy I, Inam H, Tomaszkiewicz M, Reynolds J, McCandlish D, 
            Pritchard J
  TITLE     A side-by-side comparison of variant function measurements using 
            deep mutational scanning and base editing.
  JOURNAL   Nucleic Acids Res. 2025 Jul 19;53(14):gkaf738. doi: 
            10.1093/nar/gkaf738.
  PUBMED    40744495
REFERENCE   2  (bases 1 to 12911)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 17, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res."; date: "2025-07-19"; volume: "53(14):gkaf738. doi"; 
            pages: " 10.1093/nar/gkaf738"
COMMENT     Sequence Label: nSpG UGI Puro
FEATURES             Location/Qualifiers
     source          1..12911
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1040..1161
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1051..1070)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1105..1124
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1291..1665)
                     /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"
     promoter        complement(1684..1731)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(1779..2108)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1793..1928
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1847..1866)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2093..2113
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2122..2550)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2232..2253)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2444..2463
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2596..2820
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2809..2826
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             2852..3085
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     misc_feature    3157..3745
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3263..3274
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3672..3692
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3761..4357)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3844..3864)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             complement(4358..4414)
                     /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             complement(4424..4444)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(4487..4735)
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             complement(4766..8866)
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFLWPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAKQLQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKQYRSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     promoter        complement(8904..9115)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     primer_bind     complement(9120..9140)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    9191..9308
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(9606..9650)
                     /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    9835..10068
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    10561..10686
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             10733..10913
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(10931..11129)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(10956..10976)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        11131..11510
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     11682..11701
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        11776..11880
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(11881..11949,11950..12741)
                     /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(12099..12118)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt
     1021 cgacggtata cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     1081 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     1141 ataagctgca ataaacaagt tggggtgggc gaagaactcc agcatgagat ccccgcgctg
     1201 gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc
     1261 ggcggtggaa tcgaaatctc gtagcacgtg tcagtcctgc tcctcggcca cgaagtgcac
     1321 gcagttgccg gccgggtcgc gcagggcgaa ctcccgcccc cacggctgct cgccgatctc
     1381 ggtcatggcc ggcccggagg cgtcccggaa gttcgtggac acgacctccg accactcggc
     1441 gtacagctcg tccaggccgc gcacccacac ccaggccagg gtgttgtccg gcaccacctg
     1501 gtcctggacc gcgctgatga acagggtcac gtcgtcccgg accacaccgg cgaagtcgtc
     1561 ctccacgaag tcccgggaga acccgagccg gtcggtccag aactcgaccg ctccggcgac
     1621 gtcgcgcgcg gtgagcaccg gaacggcact ggtcaacttg gccatggttt agttcctcac
     1681 cttgtcgtat tatactatgc cgatatacta tgccgatgat taattgtcaa cacgtgctga
     1741 tcagatccga aaatggatat acaagctccc gggagctttt tgcaaaagcc taggcctcca
     1801 aaaaagcctc ctcactactt ctggaatagc tcagaggcag aggcggcctc ggcctctgca
     1861 taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg cggagttagg
     1921 ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag atgcatgctt
     1981 tgcatacttc tgcctgctgg ggagcctggg gactttccac acctggttgc tgactaattg
     2041 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acaccctaac
     2101 tgacacacat tccacagaat taattcgcgt taaatttttg ttaaatcagc tcatttttta
     2161 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     2221 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     2281 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     2341 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     2401 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     2461 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     2521 ccgcgcttaa tgcgccgcta cagggcgcgt ggggataccc cctagagccc cagctggttc
     2581 tttccgcctc agaagccata gagcccaccg catccccagc atgcctgcta ttgtcttccc
     2641 aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga cacctactca
     2701 gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg gcaccttcca
     2761 gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta gaaggcacag
     2821 tcgaggctga tcagcgggtt taaacgggcc ctgctagaga ttttccacac tgactaaaag
     2881 ggtctgaggg atctctagtt accagagtca cacaacagac gggcacacac tacttgaagc
     2941 actcaaggca agctttattg aggcttaagc agtgggttcc ctagttagcc agagagctcc
     3001 caggctcaga tctggtctaa ccagagagac ccagtacaag caaaaagcag atcttgtctt
     3061 cgttgggagt gaattagccc ttccagtccc cccttttctt ttaaaaagtg gctaagatct
     3121 acagctgcct tgtaagtcat tggtcttaaa gtcgacgcgg ggaggcggcc caaagggaga
     3181 tccgactcgt ctgagggcga aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc
     3241 cgcgggaagg aaggtccgct ggattgaggg ccgaagggac gtagcagaag gacgtcccgc
     3301 gcagaatcca ggtggcaaca caggcgagca gccaaggaaa ggacgatgat ttccccgaca
     3361 acaccacgga attgtcagtg cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg
     3421 cggcgatgag ttccgccgtg gcaataggga gggggaaagc gaaagtcccg gaaaggagct
     3481 gacaggtggt ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca gtgcacacca
     3541 cgccacgttg cctgacaacg ggccacaact cctcataaag agacagcaac caggatttat
     3601 acaaggagga gaaaatgaaa gccatacggg aagcaatagc atgatacaaa ggcattaaag
     3661 cagcgtatcc acatagcgta aaaggagcaa catagttaag aataccagtc aatctttcac
     3721 aaattttgta atccagaggt tgattgtcga cttaacgcgt tcaggcaccg ggcttgcggg
     3781 tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc ggcggtgacg gtgaagccga
     3841 gccgctcgta gaaggggagg ttgcggggcg cggaggtctc caggaaggcg ggcaccccgg
     3901 cgcgctcggc cgcctccact ccggggagca cgacggcgct gcccagaccc ttgccctggt
     3961 ggtcgggcga gactccgacg gtggccagga accacgcggg ctccttgggc cggtgcggcg
     4021 ccaggaggcc ttccatctgt tgctgcgcgg ccagccggga accgctcaac tcggccatgc
     4081 gcgggccgat ctcggcgaac accgcccccg cttcgacgct ctccggcgtg gtccagaccg
     4141 ccaccgcggc gccgtcgtcc gcgacccaca ccttgccgat gtcgagcccg acgcgcgtga
     4201 ggaagagttc ttgcagctcg gtgacccgct cgatgtggcg gtccggatcg acggtgtggc
     4261 gcgtggcggg gtagtcggcg aacgcggcgg cgagggtgcg tacggccctg gggacgtcgt
     4321 cgcgggtggc gaggcgcacc gtgggcttgt actcggtcgg tccaggattc tcttcgacat
     4381 ctccggcttg tttcagcaga gagaagtttg ttgcgccagc gctgactttc ctcttcttct
     4441 tgggctcgaa ctcgctgccg tcggcggttc ttttggagcc tccgctcagc atcttgatct
     4501 tattctcgcc gttagaatcc tggatgacca gggcccaagg cttatactcg ggggcgtcag
     4561 aggtcagcag catcacattc tcatctgtgg actcgtcgta ggcggtgtgc accaggatgt
     4621 cagactctgg cttgttgccg atcacttcct ccacctcctc aggcagcatc aggatggact
     4681 cctgaatgac cagctgtttc ccagtctcct tctcaatgat gtcgctcaga ttagtgctcc
     4741 ccccgctccc gccgctcccg ccgctgtcac ctcccagctg agacaggtcg atccgtgtct
     4801 cgtacaggcc ggtgatgctc tggtggatca gggtggcgtc cagcacctct ttggtgcttc
     4861 tgtactgctt ccggtcgatg gtggtgtcaa agtacttgaa ggcggcaggg gctcccagat
     4921 tggtcagggt aaacaggtgg atgatattct cggcctgctc tctgatgggc ttatcccggt
     4981 gcttgttgta ggcggacagc actttgtcca gattagcgtc ggccaggatc actctcttgg
     5041 agaactcgct gatctgctcg atgatctcgt ccaggtagtg cttgtgctgt tccacaaaca
     5101 gctgtttctg ctcattatcc tcgggggagc ccttcagctt ctcatagtgg ctggccaggt
     5161 acaggaagtt cacatatttg gagggcaggg ccagttcgtt tcccttctgc agctgcttgg
     5221 cagaggccag cattctcttc cggccgtttt ccagctcgaa cagggagtac ttaggcagct
     5281 tgatgatcag gtcctttttc acttctttgt agcccttggc ttccagaaag tcgatgggat
     5341 tcttctcgaa gctgcttctt tccatgatgg tgatccccag cagctctttc acactcttca
     5401 gtttcttgga cttgcccttt tccactttgg ccaccaccag cacagaatag gccacggtgg
     5461 gccacaggaa gccgccgtac ttcttagggt cccagtcctt ctttctggcg atcagcttat
     5521 cgctgttcct cttgggcagg atagactctt tgctgaagcc gcctgtctgc acctcggtct
     5581 ttttcacgat attcacttgg ggcatgctca gcactttccg cacggtggca aaatcccggc
     5641 ccttatccca cacgatctcc ccggtttcgc cgtttgtctc gatcagaggc cgcttccgga
     5701 tctcgccgtt ggccagggta atctcggtct tgaaaaagtt catgatgttg ctgtagaaga
     5761 agtacttggc ggtagccttg ccgatttcct gctcgctctt ggcgatcatc ttccgcacgt
     5821 cgtacacctt gtagtcgccg tacacgaact cgctttccag cttagggtac tttttgatca
     5881 gggcggttcc cacgacggcg ttcaggtagg cgtcgtgggc gtggtggtag ttgttgatct
     5941 cgcgcacttt gtaaaactgg aaatccttcc ggaaatcgga caccagcttg gacttcaggg
     6001 tgatcacttt cacttcccgg atcagcttgt cattctcgtc gtacttagtg ttcatccggg
     6061 agtccaggat ctgtgccacg tgctttgtga tctgccgggt ttccaccagc tgtctcttga
     6121 tgaagccggc cttatccagt tcgctcaggc cgcctctctc ggccttggtc agattgtcga
     6181 actttctctg ggtaatcagc ttggcgttca gcagctgccg ccagtagttc ttcatcttct
     6241 tcacgacctc ttcggagggc acgttgtcgc tcttgccccg gttcttgtcg cttctggtca
     6301 gcaccttgtt gtcgatggag tcgtccttca gaaagctctg aggcacgata tggtccacat
     6361 cgtagtcgga cagccggttg atgtccagtt cctggtccac gtacatatcc cgcccattct
     6421 gcaggtagta caggtacagc ttctcgttct gcagctgggt gttttccacg gggtgttctt
     6481 tcaggatctg gctgcccagc tctttgatgc cctcttcgat ccgcttcatt ctctcgcggc
     6541 tgttcttctg tcccttctgg gtggtctggt tctctctggc catttcgatc acgatgttct
     6601 cgggcttgtg ccggcccatc actttcacga gctcgtccac caccttcact gtctgcagga
     6661 tgcccttctt aatggcgggg ctgccggcca gattggcaat gtgctcgtgc aggctatcgc
     6721 cctggccgga cacctgggct ttctggatgt cctctttaaa ggtcaggctg tcgtcgtgga
     6781 tcagctgcat gaagtttctg ttggcgaagc cgtcggactt caggaaatcc aggattgtct
     6841 tgccggactg cttgtcccgg atgccgttga tcagcttccg gctcagcctg ccccagccgg
     6901 tgtatctccg ccgcttcagc tgcttcatca ctttgtcgtc gaacaggtgg gcataggttt
     6961 tcagccgttc ctcgatcatc tctctgtcct caaacagtgt cagggtcagc acgatatctt
     7021 ccagaatgtc ctcgttttcc tcattgtcca ggaagtcctt gtccttgata attttcagca
     7081 gatcgtggta tgtgcccagg gaggcgttga accgatcttc cacgccggag atttccacgg
     7141 agtcgaagca ctcgattttc ttgaagtagt cctctttcag ctgcttcacg gtcactttcc
     7201 ggttggtctt gaacagcagg tccacgatgg cctttttctg ctcgccgctc aggaaggcgg
     7261 gctttctcat tccctcggtc acgtatttca ctttggtcag ctcgttatac acggtgaagt
     7321 actcgtacag caggctgtgc ttgggcagca ccttctcgtt gggcaggttc ttatcgaagt
     7381 tggtcatccg ctcgatgaag ctctgggcgg aagcgccctt gtccaccact tcctcgaagt
     7441 tccagggggt gatggtttcc tcgctctttc tggtcatcca ggcgaatctg ctgtttcccc
     7501 tggccagagg gcccacgtag taggggatgc ggaaggtcag gatcttctcg atcttttccc
     7561 ggttgtcctt caggaatggg taaaaatctt cctgccgccg cagaatggcg tgcagctctc
     7621 ccaggtggat ctggtggggg atgctgccgt tgtcgaaggt ccgctgcttc cgcagcaggt
     7681 cctctctgtt cagcttcacg agcagttcct cggtgccgtc catcttttcc aggatgggct
     7741 tgatgaactt gtagaactct tcctggctgg ctccgccgtc aatgtagccg gcgtagccgt
     7801 tcttgctctg gtcgaagaaa atctctttgt acttctcagg cagctgctgc cgcacgagag
     7861 ctttcagcag ggtcaggtcc tggtggtgct cgtcgtatct cttgatcata gaggcgctca
     7921 ggggggcctt ggtgatctcg gtgttcactc tcaggatgtc gctcagcagg atggcgtcgg
     7981 acaggttctt ggcggccaga aacaggtcgg cgtactggtc gccgatctgg gccagcaggt
     8041 tgtccaggtc gtcgtcgtag gtgtccttgc tcagctgcag tttggcatcc tcggccaggt
     8101 cgaagttgct cttgaagttg ggggtcaggc ccaggctcag ggcaatcagg tttccgaaca
     8161 ggccattctt cttctcgccg ggcagctggg cgatcagatt ttccagccgt ctgctcttgc
     8221 tcagtctggc agacaggatg gccttggcgt ccacgccgct ggcgttgatg gggttttcct
     8281 cgaacagctg gttgtaggtc tgcaccagct ggatgaacag cttgtccacg tcgctgttgt
     8341 cggggttcag gtcgccctcg atcaggaagt ggccccggaa cttgatcatg tgggccaggg
     8401 ccagatagat cagccgcagg tcggccttgt cggtgctgtc caccagtttc tttctcaggt
     8461 ggtagatggt ggggtacttc tcgtggtagg ccacctcgtc cacgatgttg ccgaagatgg
     8521 ggtgccgctc gtgcttctta tcctcttcca ccaggaagga ctcttccagt ctgtggaaga
     8581 agctgtcgtc caccttggcc atctcgttgc tgaagatctc ttgcagatag cagatccggt
     8641 tcttccgtct ggtgtatctt cttctggcgg ttctcttcag ccgggtggcc tcggctgttt
     8701 cgccgctgtc gaacagcagg gctccgatca ggttcttctt gatgctgtgc cggtcggtgt
     8761 tgcccagcac cttgaatttc ttgctgggca ccttgtactc gtcggtgatc acggcccagc
     8821 ccacagagtt ggtgccgatg gccaggccga tgctgtactt cttgtcgctt ccgtcggctg
     8881 tccgtttcat ggtggcaccg gtcctgtgtt ctggcggcaa acccgttgcg aaaaagaacg
     8941 ttcacggcga ctactgcact tatatacggt tctcccccac cctcgggaaa aaggcggagc
     9001 cagtacacga catcactttc ccagtttacc ccgcgccacc ttctctaggc accggatcaa
     9061 ttgccgaccc ctccccccaa cttctcgggg actgtgggcg atgtgcgctc tgcccactga
     9121 atcatgggaa ataggccctc ggtaccttaa ttaaccaaac tggatctctg ctgtccctgt
     9181 aataaacccg aaaattttga atttttgtaa tttgtttttg taattcttta gtttgtatgt
     9241 ctgttgctat tatgtctact attctttccc ctgcactgta ccccccaatc cccccttttc
     9301 ttttaaaatt gtggatgaat actgccattt gtctgcagaa ttggcgcacg cagtgccgat
     9361 ccgttcacta atcgaatgga tctgtctctg tctctctctc caccttcttc ttctattcct
     9421 tcgggcctgt cgggtcccct cggggttggg aggtgggtct gaaacgataa tggtgaatat
     9481 ccctgcctaa ctctattcac tatagaaagt acagcaaaaa ctattcttaa acctaccaag
     9541 cctcctacta tcattatgaa taattttata taccacagcc aatttgttat gttaaaccaa
     9601 ttccacaaac ttgcccattt atctaattcc aataattctt gttcattctt ttcttgctgg
     9661 ttttgcgatt cttcaattaa ggagtgtatt aagcttgtgt aattgttaat ttctctgtcc
     9721 cactccatcc aggtcgtgtg attccaaatc tgttccagag atttattact ccaactagca
     9781 ttccaaggca cagcagtggt gcaaatgagt tttccagagc aaccccaaat ccccaggagc
     9841 tgttgatcct ttaggtatct ttccacagcc aggattcttg cctggagctg cttgatgccc
     9901 cagactgtga gttgcaacag atgctgttgc gcctcaatag ccctcagcaa attgttctgc
     9961 tgctgcacta taccagacaa taattgtctg gcctgtaccg tcagcgtcat tgacgctgcg
    10021 cccatagtgc ttcctgctgc tcccaagaac ccaaggaaca aagctcctat tcccactgct
    10081 cttttttctc tctgcaccac tcttctcttt gccttggtgg gtgctactcc taatggttca
    10141 atttttacta ctttatattt atataattca cttctccaat tgtccctcat atctcctcct
    10201 ccaggtctga agatcagcgg ccgcttgctg tgcggtggtc ttacttttgt tttgctcttc
    10261 ctctatcttg tctaaagctt ccttggtgtc ttttatctct atcctttgat gcacacaata
    10321 gagggttgct actgtattat ataatgatct aagttcttct gatcctgtct gaagggatgg
    10381 ttgtagctgt cccagtattt gtctacagcc ttctgatgtt tctaacaggc caggattaac
    10441 tgcgaatcgt tctagctccc tgcttgccca tactatatgt tttaatttat attttttctt
    10501 tccccctggc cttaaccgaa ttttttccca tcgcgatcta attctccccc gcttaatact
    10561 gacgctctcg cacccatctc tctccttcta gcctccgcta gtcaaaattt ttggcgtact
    10621 caccagtcgc cgcccctcgc ctcttgccgt gcgcgcttca gcaagccgag tcctgcgtcg
    10681 agagagctcc tctggtttcc ctttcgcttt caagtccctg ttcgggcgcc actgctagag
    10741 attttccaca ctgactaaaa gggtctgagg gatctctagt taccagagtc acacaacaga
    10801 cgggcacaca ctacttgaag cactcaaggc aagctttatt gaggcttaag cagtgggttc
    10861 cctagttagc cagagagctc ccaggctcag atctggtcta accagagaga cccagtacag
    10921 gcaaaacgcg ctgcttatat agacctccca ccgtacacgc ctaccgccca tttgcgtcaa
    10981 tggggcggag ttgttacgac attttggaaa gtcccgttga ttttggtgcc aaaacaaact
    11041 cccattgacg tcaatggggt ggagacttgg aaatccccgt gagtcaaacc gctatccacg
    11101 cccattgatg tactgccaaa accgcatcac catggtaata gcgatgacta atacgtagat
    11161 gtactgccaa gtaggaaagt cccataaggt catgtactgg gcataatgcc aggcgggcca
    11221 tttaccgtca ttgacgtcaa tagggggcgt acttggcata tgatacactt gatgtactgc
    11281 caagtgggca gtttaccgta aatactccac ccattgacgt caatggaaag tccctattgg
    11341 cgttactatg ggaacatacg tcattattga cgtcaatggg cgggggtcgt tgggcggtca
    11401 gccaggcggg ccatttaccg taagttatgt aacgcggaac tccatatatg ggctatgaac
    11461 taatgacccc gtaattgatt actattaata actagtcaat aatcaatgtc aacgcgtata
    11521 tctggcccgt acatcgcgaa gcagcgcaaa acgcctaacc ctaagcagat tcttcatgca
    11581 attgtcggtc aagccttgcc ttgttgtagc ttaaattttg ctcgcgcact actcagcgac
    11641 ctccaacaca caagcaggga gcagatactg gcttaactat gcggcatcag agcagattgt
    11701 actgagagtg caccataggg gatcgggaga tctcccgatc cgtcgacgtc aggtggcact
    11761 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
    11821 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
    11881 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
    11941 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
    12001 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
    12061 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
    12121 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
    12181 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
    12241 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
    12301 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
    12361 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
    12421 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
    12481 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
    12541 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
    12601 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
    12661 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
    12721 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
    12781 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
    12841 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
    12901 aaaggatctt c
//
