LOCUS       sequence-447701-        9591 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCMV and pT7 Human expression plasmid for TadCBEd using SpCas9 CBE 
            enzyme with MRKSKC amino acids at positions 
            1135,1136,1218,1219,1335,T1337 with C-term BPNLS-P2A-EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9591)
  AUTHORS   Silverstein RA, Kim N, Kroell AS, Walton RT, Delano J, Butcher RM, 
            Pacesa M, Smith BK, Christie KA, Ha LL, Meis RJ, Clark AB, Spinner 
            AD, Lazzarotto CR, Li Y, Matsubara A, Urbina EO, Dahl GA, Correia 
            BE, Marks DS, Tsai SQ, Pinello L, De Ravin SS, Liu Q, Kleinstiver BP
  TITLE     Custom CRISPR-Cas9 PAM variants via scalable engineering and machine
            learning.
  JOURNAL   Nature. 2025 Apr 22. doi: 10.1038/s41586-025-09021-y.
  PUBMED    40262634
REFERENCE   2  (bases 1 to 9591)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2025 Apr 22. doi: 10.1038/s41586-025-09021-y."
COMMENT     Sequence Label: pCMV-T7-TadCBEd-SpCas9(MRKSKC)-P2A-EGFP (AHK446)
FEATURES             Location/Qualifiers
     source          1..9591
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        121..500
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        501..704
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     655..675
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     746..765
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        746..764
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             814..834
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1429..5529
                     /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
                     SILPKRNSDKLIARKKDWDPKKYGGFMRPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAKSLQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKKYCSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5560..5808
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             5839..6087
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             6130..6150
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6160..6216
                     /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             join(6217..6219,6220..6222,6223..6933)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(6217..6219,6220..6222,6223..6933)
                     /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(6523..6542)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6870..6891
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             6946..6966
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6975..6992
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7015..7032)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7021..7245
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7316..7332)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7316..7332)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7329..7351)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7340..7356
                     /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(7364..7370,7371..7388,7389..7394))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7409..7430
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7547..7564)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7718..8306)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7798..7817)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8477..9268,9269..9337))
                     /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     9100..9119
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9338..9442)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(9521..9540)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
       61 atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
      121 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
      181 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      241 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      301 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      361 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      421 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      481 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      541 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      601 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      661 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      721 agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg
      781 aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa agtctctgag
      841 gtggagtttt cccacgagta ctggatgaga catgccctga ccctggccaa gagggcacgc
      901 gatgagagga aggcccctgt gggagccgtg ctggtgctga acaatagagt gatcggcgag
      961 ggctggaaca gagccatcgg cctgcacgac ccaacagccc atgccgaaat tatcgccctg
     1021 agacagggcg gcctggtcat gcagaactac agactgattg acgccaccct gtacgtgaca
     1081 ttcgagcctt gcgtgatgtg cgccggcgcc atgatcaact ctaggatcgg ccgcgtggtg
     1141 tttggcgtga ggaacagcaa aagaggcgcc gcaggctccc tgatgaacgt gctgaactac
     1201 cccggcatga atcaccgcgt cgaaattacc gagggaatcc tggcagatga atgtgccgcc
     1261 ctgctgtgcg atttctaccg gatgcctaga caggtgttca atgctcagaa gaaggcccag
     1321 agctccatca actccggagg atctagcgga ggctcctctg gctctgagac acctggcaca
     1381 agcgagagcg caacacctga aagcagcggg ggcagcagcg gggggtcaga caagaagtac
     1441 agcatcggcc tggccatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac
     1501 aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag
     1561 aacctgatcg gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag
     1621 agaaccgcca gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc
     1681 ttcagcaacg agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc
     1741 ctggtggaag aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag
     1801 gtggcctacc acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc
     1861 accgacaagg ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg
     1921 ggccacttcc tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc
     1981 atccagctgg tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc
     2041 gtggacgcca aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg
     2101 atcgcccagc tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc
     2161 ctgggcctga cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag
     2221 ctgagcaagg acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag
     2281 tacgccgacc tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc
     2341 ctgagagtga acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac
     2401 gacgagcacc accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag
     2461 aagtacaaag agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc
     2521 ggagccagcc aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc
     2581 accgaggaac tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc
     2641 gacaacggca gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg
     2701 caggaagatt tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc
     2761 ttccgcatcc cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg
     2821 accagaaaga gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc
     2881 gcttccgccc agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag
     2941 aaggtgctgc ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc
     3001 aaagtgaaat acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa
     3061 aaggccatcg tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa
     3121 gaggactact tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat
     3181 cggttcaacg cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac
     3241 ttcctggaca atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg
     3301 tttgaggaca gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac
     3361 aaagtgatga agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag
     3421 ctgatcaacg gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc
     3481 gacggcttcg ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa
     3541 gaggacatcc agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc
     3601 aatctggccg gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac
     3661 gagctcgtga aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga
     3721 gagaaccaga ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa
     3781 gagggcatca aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag
     3841 ctgcagaacg agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac
     3901 caggaactgg acatcaaccg gctgtccgac tacgatgtgg accatatcgt gcctcagagc
     3961 tttctgaagg acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc
     4021 aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag
     4081 ctgctgaacg ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga
     4141 ggcggcctga gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg
     4201 cagatcacaa agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag
     4261 aatgacaagc tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat
     4321 ttccggaagg atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac
     4381 gacgcctacc tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa
     4441 agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc
     4501 gagcaggaaa tcggcaaggc taccgccaag tacttcttct acagcaacat catgaacttt
     4561 ttcaagaccg agattaccct ggccaacggc gagatccgga agcggcctct gatcgagaca
     4621 aacggcgaaa ccggggagat cgtgtgggat aagggccggg attttgccac cgtgcggaaa
     4681 gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc
     4741 agcaaagagt ctatcctgcc caagaggaac agcgataagc tgatcgccag aaagaaggac
     4801 tgggacccta agaagtacgg cggcttcatg agacccaccg tggcctattc tgtgctggtg
     4861 gtggccaaag tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga gctgctgggg
     4921 atcaccatca tggaaagaag cagcttcgag aagaatccca tcgactttct ggaagccaag
     4981 ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc cctgttcgag
     5041 ctggaaaacg gccggaagag aatgctggcc tctgccaaga gcctgcagaa gggaaacgaa
     5101 ctggccctgc cctccaaata tgtgaacttc ctgtacctgg ccagccacta tgagaagctg
     5161 aagggctccc ccgaggataa tgagcagaaa cagctgtttg tggaacagca caagcactac
     5221 ctggacgaga tcatcgagca gatcagcgag ttctccaaga gagtgatcct ggccgacgct
     5281 aatctggaca aagtgctgtc cgcctacaac aagcaccggg ataagcccat cagagagcag
     5341 gccgagaata tcatccacct gtttaccctg accaatctgg gagcccctgc cgccttcaag
     5401 tactttgaca ccaccatcga ccggaagaag tactgcagca ccaaagaggt gctggacgcc
     5461 accctgatcc accagagcat caccggcctg tacgagacac ggatcgacct gtctcagctg
     5521 ggaggtgaca gcggcgggag cggcgggagc ggggggagca ctaatctgag cgacatcatt
     5581 gagaaggaga ctgggaaaca gctggtcatt caggagtcca tcctgatgct gcctgaggag
     5641 gtggaggaag tgatcggcaa caagccagag tctgacatcc tggtgcacac cgcctacgac
     5701 gagtccacag atgagaatgt gatgctgctg acctctgacg cccccgagta taagccttgg
     5761 gccctggtca tccaggattc taacggcgag aataagatca agatgctgag cggaggatcc
     5821 ggaggatctg gaggcagcac caacctgtct gacatcatcg agaaggagac aggcaagcag
     5881 ctggtcatcc aggagagcat cctgatgctg cccgaagaag tcgaagaagt gatcggaaac
     5941 aagcctgaga gcgatatcct ggtccatacc gcctacgacg agagtaccga cgaaaatgtg
     6001 atgctgctga catccgacgc cccagagtat aagccctggg ctctggtcat ccaggattcc
     6061 aacggagaga acaaaatcaa aatgctgtct ggcggctcaa aaagaaccgc cgacggcagc
     6121 gaattcgagc ccaagaagaa gaggaaagtc ggaagcggag ctactaactt cagcctgctg
     6181 aagcaggctg gagacgtgga ggagaaccct ggacctatgg tgagcaaggg cgaggagctg
     6241 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     6301 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     6361 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctatgga
     6421 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     6481 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     6541 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     6601 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     6661 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     6721 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     6781 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     6841 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     6901 gggatcactc tcggcatgga cgagctgtac aagtctggtg gttctcccaa gaagaagagg
     6961 aaagtctaac cggtcatcat caccatcacc attgagttta aacccgctga tcagcctcga
     7021 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     7081 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     7141 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     7201 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     7261 gaaccagctg gggctcgata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     7321 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     7381 gcataaagtg taaagcctag ggtgcctaat gagtgagcta actcacatta attgcgttgc
     7441 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     7501 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     7561 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     7621 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     7681 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     7741 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     7801 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     7861 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     7921 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     7981 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     8041 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     8101 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     8161 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     8221 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     8281 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgaca
     8341 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     8401 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     8461 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     8521 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     8581 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     8641 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     8701 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     8761 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     8821 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     8881 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     8941 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     9001 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     9061 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     9121 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     9181 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     9241 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     9301 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     9361 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     9421 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     9481 cgggagatcg atctcccgat cccctagggt cgactctcag tacaatctgc tctgatgccg
     9541 catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag t
//
