LOCUS       sequence-388891-       13677 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human codon-optimized SpCas9-miniCGBE1 and blasticidin 
            resistance: EFS promoter-SpCas9-miniCGBE1-NLS-FLAG-P2A-BSD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13677)
  AUTHORS   Kim N, Choi S, Kim S, Song M, Seo JH, Min S, Park J, Cho SR, Kim HH
  TITLE     Deep learning models to predict the editing efficiencies and 
            outcomes of diverse base editors.
  JOURNAL   Nat Biotechnol. 2024 Mar;42(3):484-497. doi: 
            10.1038/s41587-023-01792-x
  PUBMED    37188916
REFERENCE   2  (bases 1 to 13677)
  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"; journalName: 
            "Nat Biotechnol"; date: "2024-03"; volume: "42"; issue: "3"; pages: 
            "484-497"
COMMENT     Sequence Label: SpCas9-miniCGBE1
FEATURES             Location/Qualifiers
     source          1..13677
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    417..538
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(454..473)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     508..527
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(587..603)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(587..603)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(600..622)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    611..627
                     /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(635..641,642..659,660..665))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    680..701
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(818..835)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(989..1577)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1069..1088)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1748..2539,2540..2608))
                     /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     2371..2390
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2609..2713)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2788..2807)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2979..3358
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3360..3558
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3513..3533
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             3576..3756
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    3803..3928
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4421..4654
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             4839..4883
                     /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    5181..5298
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5391..5602
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             5653..5673
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(6454..6477,6478..6480,6481..10554)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             6454..10554
                     /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
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             10597..10617
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10618..10641
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             10651..10707
                     /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             10708..11106
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    11131..11719
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(11184..11204)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(11602..11613)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(11722..11738)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11723..11739)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             12244..12424
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(12450..12467)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    12456..12680
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      12726..13154
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(12813..12832)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     13023..13044
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(13163..13183)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      13328..13462
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     13389..13408
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        13524..13571
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(13590..13677,1..287)
                     /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"
ORIGIN
        1 cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg accctgttca
       61 tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg
      121 gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct
      181 ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc gccctgcgcg
      241 acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac gtgctacgag
      301 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
      361 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
      421 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
      481 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
      541 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
      601 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
      661 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
      721 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
      781 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
      841 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
      901 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
      961 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     1021 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     1081 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     1141 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     1201 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     1261 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     1321 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     1381 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     1441 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     1501 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     1561 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     1621 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     1681 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     1741 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     1801 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     1861 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     1921 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     1981 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     2041 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     2101 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     2161 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     2221 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     2281 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     2341 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     2401 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     2461 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     2521 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     2581 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     2641 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     2701 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     2761 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     2821 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     2881 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
     2941 tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta
     3001 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
     3061 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
     3121 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
     3181 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
     3241 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
     3301 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
     3361 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
     3421 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
     3481 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
     3541 ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta gaccagatct
     3601 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     3661 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     3721 tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa
     3781 agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac
     3841 ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta
     3901 gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg
     3961 ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg
     4021 ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg
     4081 ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag aagaacttag
     4141 atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga
     4201 caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca
     4261 gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg
     4321 aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa
     4381 agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt
     4441 tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca
     4501 gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc
     4561 aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg
     4621 ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac
     4681 tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga
     4741 tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa
     4801 tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg
     4861 aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg ctgtggtata
     4921 taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt tttgctgtac
     4981 tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag acccacctcc
     5041 caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga gagagagaca
     5101 gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa
     5161 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     5221 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     5281 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt ttggttaatt
     5341 agctagctag gtcttgaaag gagtgggaat tggctccggt gcccgtcagt gggcagagcg
     5401 cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgat ccggtgccta
     5461 gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc
     5521 cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa
     5581 cgggtttgcc gccagaacac aggaccggtt gccaccatga aacggacagc cgacggaagc
     5641 gagttcgagt caccaaagaa gaagcggaaa gtctcctcag agactgggcc tgtcgccgtc
     5701 gatccaaccc tgcgccgccg gattgaacct cacgagtttg aagtgttctt tgacccccgg
     5761 gagctggcca aggagacatg cctgctgtac gagatcaact ggggaggcag gcactccatc
     5821 tggaggcaca cctctcagaa cacaaataag cacgtggagg tgaacttcat cgagaagttt
     5881 accacagagc ggtacttctg ccccaatacc agatgtagca tcacatggtt tctgagctgg
     5941 tccccttgcg gagagtgtag cagggccatc accgagttcc tgtccagata tccacacgtg
     6001 acactgttta tctacatcgc caggctgtat caccacgcag acccaaggaa taggcagggc
     6061 ctgcgcgatc tgatcagctc cggcgtgacc atccagatca tgacagagca ggagtccggc
     6121 tactgctggc ggaacttcgt gaattattct cctagcaacg aggcccactg gcctaggtac
     6181 ccacacctgt gggtgcgcct gtacgtgctg gagctgtatt gcatcatcct gggcctgccc
     6241 ccttgtctga atatcctgcg gagaaagcag ccccagctga ccttctttac aatcgccctg
     6301 cagtcttgtc actatcagag gctgccaccc cacatcctgt gggccacagg cctgaagtct
     6361 ggaggatcta gcggaggatc ctctggcagc gagacaccag gaacaagcga gtcagcaaca
     6421 ccagagagca gtggcggcag cagcggcggc agcgacaaga agtacagcat cggcctggcc
     6481 atcggcacca actctgtggg ctgggccgtg atcaccgacg agtacaaggt gcccagcaag
     6541 aaattcaagg tgctgggcaa caccgaccgg cacagcatca agaagaacct gatcggagcc
     6601 ctgctgttcg acagcggcga aacagccgag gccacccggc tgaagagaac cgccagaaga
     6661 agatacacca gacggaagaa ccggatctgc tatctgcaag agatcttcag caacgagatg
     6721 gccaaggtgg acgacagctt cttccacaga ctggaagagt ccttcctggt ggaagaggat
     6781 aagaagcacg agcggcaccc catcttcggc aacatcgtgg acgaggtggc ctaccacgag
     6841 aagtacccca ccatctacca cctgagaaag aaactggtgg acagcaccga caaggccgac
     6901 ctgcggctga tctatctggc cctggcccac atgatcaagt tccggggcca cttcctgatc
     6961 gagggcgacc tgaaccccga caacagcgac gtggacaagc tgttcatcca gctggtgcag
     7021 acctacaacc agctgttcga ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc
     7081 atcctgtctg ccagactgag caagagcaga cggctggaaa atctgatcgc ccagctgccc
     7141 ggcgagaaga agaatggcct gttcggaaac ctgattgccc tgagcctggg cctgaccccc
     7201 aacttcaaga gcaacttcga cctggccgag gatgccaaac tgcagctgag caaggacacc
     7261 tacgacgacg acctggacaa cctgctggcc cagatcggcg accagtacgc cgacctgttt
     7321 ctggccgcca agaacctgtc cgacgccatc ctgctgagcg acatcctgag agtgaacacc
     7381 gagatcacca aggcccccct gagcgcctct atgatcaaga gatacgacga gcaccaccag
     7441 gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta caaagagatt
     7501 ttcttcgacc agagcaagaa cggctacgcc ggctacattg acggcggagc cagccaggaa
     7561 gagttctaca agttcatcaa gcccatcctg gaaaagatgg acggcaccga ggaactgctc
     7621 gtgaagctga acagagagga cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc
     7681 ccccaccaga tccacctggg agagctgcac gccattctgc ggcggcagga agatttttac
     7741 ccattcctga aggacaaccg ggaaaagatc gagaagatcc tgaccttccg catcccctac
     7801 tacgtgggcc ctctggccag gggaaacagc agattcgcct ggatgaccag aaagagcgag
     7861 gaaaccatca ccccctggaa cttcgaggaa gtggtggaca agggcgcttc cgcccagagc
     7921 ttcatcgagc ggatgaccaa cttcgataag aacctgccca acgagaaggt gctgcccaag
     7981 cacagcctgc tgtacgagta cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg
     8041 accgagggaa tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc catcgtggac
     8101 ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag
     8161 aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc
     8221 ctgggcacat accacgatct gctgaaaatt atcaaggaca aggacttcct ggacaatgag
     8281 gaaaacgagg acattctgga agatatcgtg ctgaccctga cactgtttga ggacagagag
     8341 atgatcgagg aacggctgaa aacctatgcc cacctgttcg acgacaaagt gatgaagcag
     8401 ctgaagcggc ggagatacac cggctggggc aggctgagcc ggaagctgat caacggcatc
     8461 cgggacaagc agtccggcaa gacaatcctg gatttcctga agtccgacgg cttcgccaac
     8521 agaaacttca tgcagctgat ccacgacgac agcctgacct ttaaagagga catccagaaa
     8581 gcccaggtgt ccggccaggg cgatagcctg cacgagcaca ttgccaatct ggccggcagc
     8641 cccgccatta agaagggcat cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg
     8701 atgggccggc acaagcccga gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc
     8761 cagaagggac agaagaacag ccgcgagaga atgaagcgga tcgaagaggg catcaaagag
     8821 ctgggcagcc agatcctgaa agaacacccc gtggaaaaca cccagctgca gaacgagaag
     8881 ctgtacctgt actacctgca gaatgggcgg gatatgtacg tggaccagga actggacatc
     8941 aaccggctgt ccgactacga tgtggaccat atcgtgcctc agagctttct gaaggacgac
     9001 tccatcgaca acaaggtgct gaccagaagc gacaagaacc ggggcaagag cgacaacgtg
     9061 ccctccgaag aggtcgtgaa gaagatgaag aactactggc ggcagctgct gaacgccaag
     9121 ctgattaccc agagaaagtt cgacaatctg accaaggccg agagaggcgg cctgagcgaa
     9181 ctggataagg ccggcttcat caagagacag ctggtggaaa cccggcagat cacaaagcac
     9241 gtggcacaga tcctggactc ccggatgaac actaagtacg acgagaatga caagctgatc
     9301 cgggaagtga aagtgatcac cctgaagtcc aagctggtgt ccgatttccg gaaggatttc
     9361 cagttttaca aagtgcgcga gatcaacaac taccaccacg cccacgacgc ctacctgaac
     9421 gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc tggaaagcga gttcgtgtac
     9481 ggcgactaca aggtgtacga cgtgcggaag atgatcgcca agagcgagca ggaaatcggc
     9541 aaggctaccg ccaagtactt cttctacagc aacatcatga actttttcaa gaccgagatt
     9601 accctggcca acggcgagat ccggaagcgg cctctgatcg agacaaacgg cgaaaccggg
     9661 gagatcgtgt gggataaggg ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc
     9721 caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa agagtctatc
     9781 ctgcccaaga ggaacagcga taagctgatc gccagaaaga aggactggga ccctaagaag
     9841 tacggcggct tcgacagccc caccgtggcc tattctgtgc tggtggtggc caaagtggaa
     9901 aagggcaagt ccaagaaact gaagagtgtg aaagagctac tggggatcac catcatggaa
     9961 agaagcagct tcgagaagaa tcccatcgac tttctggaag ccaagggcta caaagaagtg
    10021 aaaaaggacc tgatcatcaa gctgcctaag tactccctgt tcgagctgga aaacggccgg
    10081 aagagaatgc tggcctctgc cggcgaactg cagaagggaa acgaactggc cctgccctcc
    10141 aaatatgtga acttcctgta cctggccagc cactatgaga agctgaaggg ctcccccgag
    10201 gataatgagc agaaacagct gtttgtggaa cagcacaagc actacctgga cgagatcatc
    10261 gagcagatca gcgagttctc caagagagtg atcctggccg acgctaatct ggacaaagtg
    10321 ctgtccgcct acaacaagca ccgggataag cccatcagag agcaggccga gaatatcatc
    10381 cacctgttta ccctgaccaa tctgggagcc cctgccgcct tcaagtactt tgacaccacc
    10441 atcgaccgga agaggtacac cagcaccaaa gaggtgctgg acgccaccct gatccaccag
    10501 agcatcaccg gcctgtacga gacacggatc gacctgtctc agctgggagg cgactctggc
    10561 ggctcaaaaa gaaccgccga cggcagcgaa ttcgagccca agaagaagag gaaagtcgat
    10621 tacaaagacg atgacgataa gggatccggc gcaacaaact tctctctgct gaaacaagcc
    10681 ggagatgtcg aagagaatcc tggaccgatg gccaagcctt tgtctcaaga agaatccacc
    10741 ctcattgaaa gagcaacggc tacaatcaac agcatcccca tctctgaaga ctacagcgtc
    10801 gccagcgcag ctctctctag cgacggccgc atcttcactg gtgtcaatgt atatcatttt
    10861 actgggggac cttgtgcaga actcgtggtg ctgggcactg ctgctgctgc ggcagctggc
    10921 aacctgactt gtatcgtcgc gatcggaaat gagaacaggg gcatcttgag cccctgcgga
    10981 cggtgccgac aggtgcttct cgatctgcat cctgggatca aagccatagt gaaggacagt
    11041 gatggacagc cgacggcagt tgggattcgt gaattgctgc cctctggtta tgtgtgggag
    11101 ggctaagaat tcgatatcaa gcttatcggt aatcaacctc tggattacaa aatttgtgaa
    11161 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
    11221 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
    11281 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
    11341 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
    11401 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
    11461 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
    11521 gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg
    11581 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
    11641 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
    11701 ctttgggccg cctccccgca tcgataccgt cgacctcgag acctagaaaa acatggagca
    11761 atcacaagta gcaatacagc agctaccaat gctgattgtg cctggctaga agcacaagag
    11821 gaggaggagg tgggttttcc agtcacacct caggtacctt taagaccaat gacttacaag
    11881 gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac
    11941 tcccaacgaa gacaagatat ccttgatctg tggatctacc acacacaagg ctacttccct
    12001 gattggcaga actacacacc agggccaggg atcagatatc cactgacctt tggatggtgc
    12061 tacaagctag taccagttga gcaagagaag gtagaagaag ccaatgaagg agagaacacc
    12121 cgcttgttac accctgtgag cctgcatggg atggatgacc cggagagaga agtattagag
    12181 tggaggtttg acagccgcct agcatttcat cacatggccc gagagctgca tccggactgt
    12241 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
    12301 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
    12361 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
    12421 agcagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
    12481 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
    12541 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
    12601 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
    12661 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
    12721 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
    12781 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
    12841 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
    12901 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
    12961 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
    13021 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
    13081 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
    13141 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
    13201 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
    13261 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
    13321 accatagtcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
    13381 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
    13441 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
    13501 attttcggat ctgatcagca cgtgttgaca attaatcatc ggcatagtat atcggcatag
    13561 tataatacga caaggtgagg aactaaacca tggccaagtt gaccagtgcc gttccggtgc
    13621 tcaccgcgcg cgacgtcgcc ggagcggtcg agttctggac cgaccggctc gggttct
//
