LOCUS       sequence-458313-       14794 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ef1a-AID*-dCas9-T2A-tagBFP-wPRE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14794)
  AUTHORS   Schwartz CI, Abell NS, Li A, Aradhana, Tycko J, Truong A, Montgomery
            SB, Hess GT
  TITLE     Toward optimizing diversifying base editors for high-throughput 
            mutational scanning studies.
  JOURNAL   Nucleic Acids Res. 2025 Jun 20;53(12):gkaf620. doi: 
            10.1093/nar/gkaf620.
  PUBMED    40613705
REFERENCE   2  (bases 1 to 14794)
  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: "Nucleic 
            Acids Res."; date: "2025-06-20"; volume: "53(12):gkaf620. doi"; 
            pages: " 10.1093/nar/gkaf620"
COMMENT     Sequence Label: pGH484
FEATURES             Location/Qualifiers
     source          1..14794
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2687..3865
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2918..3856
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3813..3833
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      3872..3881
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4511..4534,4535..4537,4538..7093,7094..7096,
                     7097..8611)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             8666..8686
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8702..8755
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             8756..9454
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     misc_feature    9486..10074
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9539..9559)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9957..9968)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(10077..10093)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10078..10094)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             10599..10779
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10805..10822)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10811..11035
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      11081..11509
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11168..11187)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11378..11399
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(11518..11538)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        11523..11852
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      11703..11838
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11765..11784
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11900..11947
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11966..12340
                     /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"
     polyA_signal    12470..12591
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12507..12526)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     12561..12580
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(12640..12656)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12640..12656)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12653..12675)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12664..12680
                     /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(12688..12694,12695..12712,12713..12718))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12733..12754
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12871..12888)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(13042..13630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(13122..13141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13801..14592,14593..14661))
                     /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     14424..14443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14662..14766)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta gctagctgca aagatggata aagttttaaa cagagaggaa
     2641 tctttgcagc taatggacct tctaggtctt gaaaggagtg ggaattggct ccggtgcccg
     2701 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
     2761 ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
     2821 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
     2881 cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg
     2941 cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct
     3001 gcagtacgtg attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc
     3061 ttgcgcttaa ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg
     3121 ccgccgcgtg cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct
     3181 agccatttaa aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt
     3241 aaatgcgggc caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg
     3301 ggcccgtgcg tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag
     3361 aatcggacgg gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc
     3421 gtgtatcgcc ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga
     3481 aagatggccg cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg
     3541 agagcgggcg ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc
     3601 ttcatgtgac tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt
     3661 ttggagtacg tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac
     3721 tgagtgggtg gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt
     3781 tgcccttttt gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt
     3841 ttttcttcca tttcaggtgt cgtgacgtac ggccaccatg gacagcctct tgatgaaccg
     3901 gagggagttt ctttaccaat tcaaaaatgt ccgctgggct aagggtcggc gtgagaccta
     3961 cctgtgctac gtagtgaaga ggcgtgacag tgctacatcc ttttcactgg actttggtta
     4021 tcttcgcaat aagaacggct gccacgtgga attgctcttc ctccgctaca tctcggactg
     4081 ggacctagac cctggccgct gctaccgcgt cacctggttc atctcctgga gcccctgcta
     4141 cgactgtgcc cgacatgtgg ccgactttct gcgagggaac cccaacctca gtctgaggat
     4201 cttcaccgcg cgcctctact tctgtgagga ccgcaaggct gagcccgagg ggctgcggcg
     4261 gctgcaccgc gccggggtgc aaatagccat catgaccttc aaagattatt tttactgctg
     4321 gaatactttt gtagaaaacc acggaagaac tttcaaagcc tgggaagggc tgcatgaaaa
     4381 ttcagttcgt ctctccagac agcttcggcg catccttttg cccctgtatg aggttgatga
     4441 cttacgagac gcatttcgta ctagcggcag cgagactccc gggacctcag agtccgccac
     4501 acccgaaagt gacaagaagt acagcatcgg cctggccatc ggcaccaact ctgtgggctg
     4561 ggccgtgatc accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac
     4621 cgaccggcac agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac
     4681 agccgaggcc acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg
     4741 gatctgctat ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt
     4801 ccacagactg gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat
     4861 cttcggcaac atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct
     4921 gagaaagaaa ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct
     4981 ggcccacatg atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa
     5041 cagcgacgtg gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga
     5101 aaaccccatc aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa
     5161 gagcagacgg ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt
     5221 cggcaacctg attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct
     5281 ggccgaggat gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct
     5341 gctggcccag atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga
     5401 cgccatcctg ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag
     5461 cgcctctatg atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct
     5521 cgtgcggcag cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg
     5581 ctacgccggc tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc
     5641 catcctggaa aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct
     5701 gctgcggaag cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga
     5761 gctgcacgcc attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga
     5821 aaagatcgag aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg
     5881 aaacagcaga ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt
     5941 cgaggaagtg gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt
     6001 cgataagaac ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt
     6061 caccgtgtat aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc
     6121 cttcctgagc ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa
     6181 agtgaccgtg aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt
     6241 ggaaatctcc ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct
     6301 gaaaattatc aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga
     6361 tatcgtgctg accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac
     6421 ctatgcccac ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg
     6481 ctggggcagg ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac
     6541 aatcctggat ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca
     6601 cgacgacagc ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga
     6661 tagcctgcac gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct
     6721 gcagacagtg aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa
     6781 catcgtgatc gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg
     6841 cgagagaatg aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga
     6901 acaccccgtg gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa
     6961 tgggcgggat atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt
     7021 ggaccacatc gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac
     7081 cagaagcgac aaggcccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa
     7141 gatgaagaac tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga
     7201 caatctgacc aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa
     7261 gagacagctg gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg
     7321 gatgaacact aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct
     7381 gaagtccaag ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat
     7441 caacaactac caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat
     7501 caaaaagtac cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt
     7561 gcggaagatg atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt
     7621 ctacagcaac atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg
     7681 gaagcggcct ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg
     7741 ggattttgcc accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac
     7801 cgaggtgcag acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa
     7861 gctgatcgcc agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac
     7921 cgtggcctat tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa
     7981 gagtgtgaaa gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc
     8041 catcgacttt ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct
     8101 gcctaagtac tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg
     8161 cgaactgcag aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct
     8221 ggccagccac tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt
     8281 tgtggaacag cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa
     8341 gagagtgatc ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg
     8401 ggataagccc atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct
     8461 gggagcccct gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag
     8521 caccaaagag gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac
     8581 acggatcgac ctgtctcagc tgggaggcga cagcgctgga ggaggtggaa gcggaggagg
     8641 aggaagcgga ggaggaggta gcggacctaa gaaaaagagg aaggtggcgg ccgctggatc
     8701 cgagggcaga ggaagtctgc taacatgcgg tgacgtcgag gagaatcctg gcccaagcga
     8761 gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtgg acaaccatca
     8821 cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga ccatgagaat
     8881 caaggtggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta ctagcttcct
     8941 ctacggcagc aagaccttca tcaaccacac ccagggcatc cccgacttct tcaagcagtc
     9001 cttccctgag ggcttcacat gggagagagt caccacatac gaagacgggg gcgtgctgac
     9061 cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca agatcagagg
     9121 ggtgaacttc acatccaacg gccctgtgat gcagaagaaa acactcggct gggaggcctt
     9181 caccgagacg ctgtaccccg ctgacggcgg cctggaaggc agaaacgaca tggccctgaa
     9241 gctcgtgggc gggagccatc tgatcgcaaa catcaagacc acatatagat ccaagaaacc
     9301 cgctaagaac ctcaagatgc ctggcgtcta ctatgtggac tacagactgg aaagaatcaa
     9361 ggaggccaac aacgagacct acgtcgagca gcacgaggtg gcagtggcca gatactgcga
     9421 cctccctagc aaactggggc acaagcttaa ttagtaaggc cgaattcgat atcaagctta
     9481 tcggtaatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     9541 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     9601 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     9661 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     9721 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     9781 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     9841 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     9901 tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     9961 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
    10021 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat
    10081 accgtcgacc tcgagaccta gaaaaacatg gagcaatcac aagtagcaat acagcagcta
    10141 ccaatgctga ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt tttccagtca
    10201 cacctcaggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
    10261 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa gatatccttg
    10321 atctgtggat ctaccacaca caaggctact tccctgattg gcagaactac acaccagggc
    10381 cagggatcag atatccactg acctttggat ggtgctacaa gctagtacca gttgagcaag
    10441 agaaggtaga agaagccaat gaaggagaga acacccgctt gttacaccct gtgagcctgc
    10501 atgggatgga tgacccggag agagaagtat tagagtggag gtttgacagc cgcctagcat
    10561 ttcatcacat ggcccgagag ctgcatccgg actgtactgg gtctctctgg ttagaccaga
    10621 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
    10681 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
    10741 ccctcagacc cttttagtca gtgtggaaaa tctctagcag ggcccgttta aacccgctga
    10801 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
    10861 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
    10921 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
    10981 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
    11041 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
    11101 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
    11161 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
    11221 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
    11281 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
    11341 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
    11401 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
    11461 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
    11521 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
    11581 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
    11641 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
    11701 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
    11761 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
    11821 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
    11881 tcggatctga tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata
    11941 atacgacaag gtgaggaact aaaccatggc caagttgacc agtgccgttc cggtgctcac
    12001 cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga
    12061 cttcgtggag gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc
    12121 ggtccaggac caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga
    12181 cgagctgtac gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc
    12241 ggccatgacc gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc
    12301 cggcaactgc gtgcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga
    12361 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
    12421 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
    12481 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
    12541 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
    12601 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
    12661 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
    12721 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
    12781 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
    12841 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
    12901 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
    12961 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
    13021 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
    13081 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
    13141 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
    13201 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
    13261 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
    13321 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
    13381 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
    13441 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
    13501 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
    13561 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
    13621 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
    13681 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
    13741 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
    13801 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
    13861 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
    13921 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
    13981 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
    14041 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
    14101 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
    14161 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
    14221 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
    14281 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
    14341 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
    14401 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
    14461 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
    14521 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
    14581 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
    14641 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
    14701 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
    14761 tccgcgcaca tttccccgaa aagtgccacc tgac
//
