LOCUS       Exported                9912 bp ds-DNA     circular SYN 21-MAR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9912)
  AUTHORS   NIH-NCI
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Mar 25, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9912
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     1..19
                     /label=A192-px459v2-deltaSpCas9-FP
     primer_bind     1..19
                     /label=A219-px459v2-dSpCas9-XhoI-F
     CDS             16..69
                     /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             70..669
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(70..89)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     566..586
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    668..669
                     /label=Sanger sequencing confirmed silent mutation
     primer_bind     complement(697..714)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    703..910
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   919..1059
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   919..1048
                     /label=AAV2 ITR
     rep_origin      1134..1589
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1221..1240)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1431..1452
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1606..1625)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1725..1747
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1785..1803)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1845..1862)
                     /label=pQE promoter
     promoter        1871..1975
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1976..2836
                     /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(2194..2213)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3007..3595
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3228..3247
                     /label=RepO-R
     primer_bind     complement(3354..3372)
                     /label=RepO-F
     primer_bind     3496..3515
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3657..3897
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     3657..3677
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     3701..3725
                     /label=TRC-F
     primer_bind     3828..3847
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        3924..3999
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        4096..4381
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        4383..4660
                     /label=chicken beta-actin promoter
     intron          4661..4888
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      4900..4909
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             4909..4974
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     4962..4999
                     /label=A177-px461-ABE8e-SpCas9-FP
     primer_bind     complement(4962..4980)
                     /label=A220-px459v2-dSpCas9-XhoI-R
     primer_bind     complement(4962..4980)
                     /label=A193-px459v2-deltaSpCas9-RP
     CDS             4962..4974
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DDDK"
     primer_bind     4978..5002
                     /label=EDb 3.1_F
     misc_feature    4981..5472
                     /label=TadA8e
     primer_bind     4981..5003
                     /label=ABE8e_F
     primer_bind     4981..4999
                     /label=Primer 1
     primer_bind     complement(5453..5472)
                     /label=ABE8e_R
     primer_bind     complement(5559..5580)
                     /label=Fragment.REV
     primer_bind     complement(5559..5574)
                     /label=Infusion fragment R
     primer_bind     complement(5564..5595)
                     /label=ED 3.1_R
     primer_bind     complement(5567..5583)
                     /label=EDb 6.1-2_R
     primer_bind     5578..5605
                     /label=EDb 1.0/3.0_F
     CDS             5584..9684
                     /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"
     primer_bind     complement(5639..5659)
                     /label=Cas9 rev seq1
     primer_bind     5952..5971
                     /label=Cas9_confirm_2
     primer_bind     6552..6571
                     /label=Cas9_confirm_3
     primer_bind     7152..7171
                     /label=Cas9_confirm_4
     primer_bind     7732..7751
                     /label=Cas9_5_F
     primer_bind     7732..7751
                     /label=Cas9_5_F (1)
     primer_bind     7752..7767
                     /label=Cas9_confirm_5
     primer_bind     7935..7954
                     /label=Cas9 intern seq 3
     primer_bind     8352..8371
                     /label=Cas9_confirm_6
     primer_bind     8952..8974
                     /label=Cas9_confirm_7
     primer_bind     9552..9576
                     /label=Cas9_comfirm_8
     primer_bind     complement(9678..9715)
                     /label=EDb 1.0_R
     primer_bind     complement(9680..9715)
                     /label=ED 3.0_R
     primer_bind     complement(9684..9693)
                     /label=Infusion fragment R
     primer_bind     complement(9684..9693)
                     /label=Fragment.REV
     primer_bind     9701..9723
                     /label=inf 6nls.FOR
     primer_bind     complement(9701..9717)
                     /label=Infusion vec R
     CDS             9706..9732
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             9742..9768
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             9778..9804
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             9814..9840
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             9850..9876
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             9886..9912
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     primer_bind     complement(9892..9912)
                     /label=EDi 1-3_R
     primer_bind     complement(join(9893..9912,1..19))
                     /label=A180-px461-BEACON2-SpCas9-RP
     primer_bind     complement(join(9894..9912,1))
                     /label=inf 6nls.REV
     primer_bind     complement(9894..9912)
                     /label=Primer 2
ORIGIN
        1 gaattcggca gtggagaggg cagaggaagt ctgctaacat gcggtgacgt cgaggagaat
       61 cctggcccaa tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
      121 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
      181 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
      241 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
      301 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
      361 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
      421 cccaaggagc ccgcgtggtt cctggccacc gtcggagtct cgcccgacca ccagggcaag
      481 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
      541 gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
      601 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
      661 ggtgcctagg aattctaact agagctcgct gatcagcctc gactgtgcct tctagttgcc
      721 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
      781 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
      841 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagag aatagcaggc
      901 atgctgggga gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
      961 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
     1021 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc
     1081 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
     1141 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     1201 tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     1261 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     1321 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     1381 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     1441 gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat
     1501 tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     1561 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
     1621 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     1681 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     1741 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     1801 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     1861 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     1921 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     1981 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     2041 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     2101 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     2161 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     2221 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     2281 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     2341 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     2401 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     2461 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     2521 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     2581 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     2641 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg
     2701 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     2761 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     2821 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     2881 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     2941 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     3001 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     3061 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     3121 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     3181 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     3241 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3301 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3361 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3421 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3481 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3541 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3601 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtgagg
     3661 gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt agagagataa
     3721 ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga cgtagaaagt
     3781 aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac tatcatatgc
     3841 ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg aaaggacgaa
     3901 acaccgggtc ttcgagaaga cctgttttag agctagaaat agcaagttaa aataaggcta
     3961 gtccgttatc aacttgaaaa agtggcaccg agtcggtgct tttttgtttt agagctagaa
     4021 atagcaagtt aaaataaggc tagtccgttt ttagcgcgtg cgccaattct gcagacaaat
     4081 ggctctagag gtacccgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     4141 cgacccccgc ccattgacgt caatagtaac gccaataggg actttccatt gacgtcaatg
     4201 ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag
     4261 tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattgtg cccagtacat
     4321 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
     4381 ggtcgaggtg agccccacgt tctgcttcac tctccccatc tcccccccct ccccaccccc
     4441 aattttgtat ttatttattt tttaattatt ttgtgcagcg atgggggcgg gggggggggg
     4501 ggggcgcgcg ccaggcgggg cggggcgggg cgaggggcgg ggcggggcga ggcggagagg
     4561 tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg
     4621 gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgcgctgcct
     4681 tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg ccccggctct gactgaccgc
     4741 gttactccca caggtgagcg ggcgggacgg cccttctcct ccgggctgta attagctgag
     4801 caagaggtaa gggtttaagg gatggttggt tggtggggta ttaatgttta attacctgga
     4861 gcacctgcct gaaatcactt tttttcaggt tggaccggtg ccaccatgga ctataaggac
     4921 cacgacggag actacaagga tcatgatatt gattacaaag acgatgacga taagatggcc
     4981 tctgaggtgg agttttccca cgagtactgg atgagacatg ccctgaccct ggccaagagg
     5041 gcacgggatg agagggaggt gcctgtggga gccgtgctgg tgctgaacaa tagagtgatc
     5101 ggcgagggct ggaacagagc catcggcctg cacgacccaa cagcccatgc cgaaattatg
     5161 gccctgagac agggcggcct ggtcatgcag aactacagac tgattgacgc caccctgtac
     5221 gtgacattcg agccttgcgt gatgtgcgcc ggcgccatga tccactctag gatcggccgc
     5281 gtggtgtttg gcgtgaggaa ctcaaaaaga ggcgccgcag gctccctgat gaacgtgctg
     5341 aactaccccg gcatgaatca ccgcgtcgaa attaccgagg gaatcctggc agatgaatgt
     5401 gccgccctgc tgtgcgattt ctatcggatg cctagacagg tgttcaatgc tcagaagaag
     5461 gcccagagct ccatcaactc cggaggatct agcggaggat cctctggcag cgagacacca
     5521 ggaacaagcg agtcagcaac accagagagc agtggcggca gcagcggggg gtcagctagc
     5581 gctgacaaga agtacagcat cggcctggcc atcggcacca actctgtggg ctgggccgtg
     5641 atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa caccgaccgg
     5701 cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga aacagccgag
     5761 gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa ccggatctgc
     5821 tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt cttccacaga
     5881 ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc catcttcggc
     5941 aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca cctgagaaag
     6001 aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc cctggcccac
     6061 atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga caacagcgac
     6121 gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga ggaaaacccc
     6181 atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag caagagcaga
     6241 cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct gttcggaaac
     6301 ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga cctggccgag
     6361 gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa cctgctggcc
     6421 cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc cgacgccatc
     6481 ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct gagcgcctct
     6541 atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg
     6601 cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa cggctacgcc
     6661 ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa gcccatcctg
     6721 gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg
     6781 aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg agagctgcac
     6841 gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg ggaaaagatc
     6901 gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc
     6961 agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa
     7021 gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag
     7081 aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg
     7141 tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg
     7201 agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc
     7261 gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc
     7321 tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt
     7381 atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg
     7441 ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc
     7501 cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac cggctggggc
     7561 aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg
     7621 gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac
     7681 agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg
     7741 cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca
     7801 gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga gaacatcgtg
     7861 atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga
     7921 atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc
     7981 gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg
     8041 gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga tgtggaccat
     8101 atcgtgcctc agagctttct gaaggacgac tccatcgaca acaaggtgct gaccagaagc
     8161 gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag
     8221 aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg
     8281 accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag
     8341 ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac
     8401 actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc
     8461 aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac
     8521 taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag
     8581 taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag
     8641 atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc
     8701 aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat ccggaagcgg
     8761 cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt
     8821 gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg
     8881 cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga taagctgatc
     8941 gccagaaaga aggactggga ccctaagaag tacggcggct tcgacagccc caccgtggcc
     9001 tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact gaagagtgtg
     9061 aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa tcccatcgac
     9121 tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag
     9181 tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc cggcgaactg
     9241 cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta cctggccagc
     9301 cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct gtttgtggaa
     9361 cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc caagagagtg
     9421 atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca ccgggataag
     9481 cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa tctgggagcc
     9541 cctgccgcct tcaagtactt tgacaccacc atcgaccgga agaggtacac cagcaccaaa
     9601 gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga gacacggatc
     9661 gacctgtctc agctgggagg tgacagcggc gggaattcca gatcccctgc tgctaaacgt
     9721 gttaagcttg atgggggtag cccggcagcc aagagagtca aactcgacgg cggctcccca
     9781 gcggcaaaaa gggtgaaact agacgggggt agccccgccg cgaagcgtgt aaagctggat
     9841 ggaggatcgc ctgcggctaa gcgagtcaaa ttagacggtg ggtcaccggc tgcaaagcgc
     9901 gtcaaattgg at
//
