LOCUS       sequence-256337-       11406 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac vector for CRISPR/SpCas9 applications (nuclease, base 
            editing, or epigenetic modifications). Provides gRNA and EspCas9 
            expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11406)
  AUTHORS   Fischer AD, Veronese-Paniagua DA, Swaminathan S, Kashima H, Rubin 
            DC, Madison BB
  TITLE     The oncogenic function of PLAGL2 is mediated via ASCL2 and IGF2 and 
            a Wnt-independent mechanism in colorectal cancer.
  JOURNAL   Am J Physiol Gastrointest Liver Physiol. 2023 Jun 13. doi: 
            10.1152/ajpgi.00058.2022.
  PUBMED    37310750
REFERENCE   2  (bases 1 to 11406)
  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: "Am J 
            Physiol Gastrointest Liver Physiol. 2023 Jun 13. doi: 
            10.1152/ajpgi.00058.2022."
COMMENT     Sequence Label: BII-gR-BtW-eSpCas9
FEATURES             Location/Qualifiers
     source          1..11406
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   5..39
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(308..324)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(308..324)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        580..828
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     580..600
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     751..770
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        849..924
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      1638..1647
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1644..2039
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             2058..2111
                     /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             2121..2144
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2163..2183
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(2208..4745,4746..4748,4749..5210,5211..5213,5214..5381,
                     5382..5384,5385..6308)
                     /codon_start=1
                     /product="Cas9 endonuclease from the Streptococcus pyogenes
                     Type II CRISPR/Cas system, mutated to improve targeting 
                     specificity (Slaymaker et al., 2016)"
                     /label=eSpCas9(1.1)
                     /note="carries the mutations K848A, K1003A, and R1060A"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLADDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPALESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKAPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             6309..6356
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     misc_feature    6383..6971
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6436..6456)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6854..6865)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    7051..7185
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7075..7094)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7129..7148
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   7963..8025
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(8155..8172)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8155..8171)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8164..8186)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8313..8768
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8400..8419)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8610..8631
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        8794..8898
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8899..8967,8968..9759)
                     /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(9117..9136)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9930..10518
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10419..10438
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     10672..10689
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    10806..10827
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(10842..10847,10848..10865,10866..10872)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10880..10896
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     10885..10907
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     10904..10920
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     10904..10920
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttaaccctag aaagatagtc tgcgtaaaat tgacgcatgc attcttgaaa tattgctctc
       61 tctttctaaa tagcgcgaat ccgtcgctgt gcatttagga catctcagtc gccgcttgga
      121 gctcccgtga ggcgtgcttg tcaatgcggt aagtgtcact gattttgaac tataacgacc
      181 gcgtgagtca aaatgacgca tgattatctt ttacgtgact tttaagattt aactcatacg
      241 ataattatat tgttatttca tgttctactt acgtgataac ttattatata tatattttct
      301 tgttatagat ccactagttc tagagggaca gccccccccc aaagccccca gggatgtaat
      361 tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct ccggtccggc
      421 gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg gggaaggtgg
      481 cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag acacctgggg
      541 ggatacgggg aaaaggcctc cagggccgcc cccttcaccg agggcctatt tcccatgatt
      601 ccttcatatt tgcatatacg atacaaggct gttagagaga taattggaat taatttgact
      661 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
      721 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag
      781 tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccgg agacgctgac
      841 ccgtctctgt tttagagcta gaaatagcaa gttaaaataa ggctagtccg ttatcaactt
      901 gaaaaagtgg caccgagtcg gtgctttttt gtttttgcct ccaaggcctg caggagtcaa
      961 tgggaaaaac ccattggagc caagtacact gactcaatag ggactttcca ttgggttttg
     1021 cccagtacat aaggtcaata gggggtgagt caacaggaaa gtcccattgg agccaagtac
     1081 attgagtcaa tagggacttt ccaatgggtt ttgcccagta cataaggtca atgggaggta
     1141 agccaatggg tttttcccat tactgacatg tatactgagt cattagggac tttccaatgg
     1201 gttttgccca gtacataagg tcaatagggg tgaatcaaca ggaaagtccc attggagcca
     1261 agtacactga gtcaataggg actttccatt gggttttgcc cagtacaaaa ggtcaatagg
     1321 gggtgagtca atgggttttt cccattattg gcacatacat aaggtcaata ggggtgacta
     1381 gtggagaaga gcatgcttga gggctgagtg cccctcagtg ggcagagagc acatggccca
     1441 cagtccctga gaagttgggg ggaggggtgg gcaattgaac tggtgcctag agaaggtggg
     1501 gcttgggtaa actgggaaag tgatgtggtg tactggctcc acctttttcc ccagggtggg
     1561 ggagaaccat atataagtgc agtagtctct gtgaacattc ggtttagtga acctgcaggt
     1621 cggccgccac gaccggtgcc accatggcca agcctttgtc tcaagaagaa tccaccctca
     1681 ttgaaagagc aacggctaca atcaacagca tccccatctc tgaagactac agcgtcgcca
     1741 gcgcagctct ctctagcgac ggccgcatct tcactggtgt caatgtatat cattttactg
     1801 ggggaccttg tgcagaactc gtggtgctgg gcactgctgc tgctgcggca gctggcaacc
     1861 tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat cttgagcccc tgcggacggt
     1921 gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc catagtgaag gacagtgatg
     1981 gacagccgac ggcagttggg attcgtgaat tgctgccctc tggttatgtg tgggagggcg
     2041 ctagccttgg ctccggcgag ggcaggggaa gtcttctaac atgcggggac gtggaggaaa
     2101 atcccggccc actcggcatg gactacaagg acgacgatga caagctcgat agcggcatgg
     2161 ccccaaagaa gaagcggaag gtcggtatcc acggagtccc agcagccgac aagaagtaca
     2221 gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc gacgagtaca
     2281 aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc atcaagaaga
     2341 acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga
     2401 gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg caagagatct
     2461 tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa gagtccttcc
     2521 tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc gtggacgagg
     2581 tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg gtggacagca
     2641 ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc aagttccggg
     2701 gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac aagctgttca
     2761 tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg
     2821 tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg gaaaatctga
     2881 tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt gccctgagcc
     2941 tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc aaactgcagc
     3001 tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc ggcgaccagt
     3061 acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg agcgacatcc
     3121 tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc aagagatacg
     3181 acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag ctgcctgaga
     3241 agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac attgacggcg
     3301 gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag atggacggca
     3361 ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag cggaccttcg
     3421 acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt ctgcggcggc
     3481 aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag atcctgacct
     3541 tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc gcctggatga
     3601 ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg gacaagggcg
     3661 cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg cccaacgaga
     3721 aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac gagctgacca
     3781 aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa
     3841 aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag cagctgaaag
     3901 aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc gtggaagatc
     3961 ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag gacaaggact
     4021 tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc ctgacactgt
     4081 ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca
     4141 aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg agccggaagc
     4201 tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc ctgaagtccg
     4261 acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg acctttaaag
     4321 aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag cacattgcca
     4381 atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag gtggtggacg
     4441 agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa atggccagag
     4501 agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag cggatcgaag
     4561 agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa aacacccagc
     4621 tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg tacgtggacc
     4681 aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg cctcagagct
     4741 ttctggccga cgactccatc gacaacaagg tgctgaccag aagcgacaag aaccggggca
     4801 agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac tggcggcagc
     4861 tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag gccgagagag
     4921 gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg gaaacccggc
     4981 agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag tacgacgaga
     5041 atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt
     5101 tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac cacgcccacg
     5161 acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct gcgctggaaa
     5221 gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg
     5281 agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc atgaactttt
     5341 tcaagaccga gattaccctg gccaacggcg agatccggaa ggcgcctctg atcgagacaa
     5401 acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc gtgcggaaag
     5461 tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca ggcggcttca
     5521 gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga aagaaggact
     5581 gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct gtgctggtgg
     5641 tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag ctgctgggga
     5701 tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg gaagccaagg
     5761 gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc ctgttcgagc
     5821 tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag ggaaacgaac
     5881 tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat gagaagctga
     5941 agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac aagcactacc
     6001 tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg gccgacgcta
     6061 atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc agagagcagg
     6121 ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc gccttcaagt
     6181 actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg ctggacgcca
     6241 ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg tctcagctgg
     6301 gaggcgacaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaagtaag
     6361 agctagagtt cgcgtctgga acaatcaacc tctggattac aaaatttgtg aaagattgac
     6421 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     6481 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     6541 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     6601 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     6661 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     6721 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaagct
     6781 gacgtccttt ccatggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     6841 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     6901 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     6961 cgcctccccg cctggaatta attctggcta agcacttcgt ggccgtcgat cgtttaaagg
     7021 gaggtagtga gtcgagatat cgatactagt gatccagaca tgataagata cattgatgag
     7081 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     7141 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     7201 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
     7261 ctctacaaat gtggtatggc tgattatgat cctctagagg gacagccccc ccccaaagcc
     7321 cccagggatg taattacgtc cctcccccgc tagggggcag cagcgagccg cccggggctc
     7381 cgctccggtc cggcgctccc cccgcatccc cgagccggca gcgtgcgggg acagcccggg
     7441 cacggggaag gtggcacggg atcgctttcc tctgaacgct tctcgctgct ctttgagcct
     7501 gcagacacct ggggggatac ggggaaaagc cataccaatg gagctcacgg ggacagcccc
     7561 cccccaaagc ccccagggat gtaattacgt ccctcccccg ctagggggca gcagcgagcc
     7621 gcccggggct ccgctccggt ccggcgctcc ccccgcatcc ccgagccggc agcgtgcggg
     7681 gacagcccgg gcacggggaa ggtggcacgg gatcgctttc ctctgaacgc ttctcgctgc
     7741 tctttgagcc tgcagacacc tggggggata cggggaaaaa gaattcgata aaagttttgt
     7801 tactttatag aagaaatttt gagtttttgt tttttttaat aaataaataa acataaataa
     7861 attgtttgtt gaatttatta ttagtatgta agtgtaaata taataaaact taatatctat
     7921 tcaaattaat aaataaacct cgatatacag accgataaaa cacatgcgtc aattttacac
     7981 atgattatct ttaacgtacg tcacaatatg attatctttc tagggttaat ctagctgcgt
     8041 gttctgcagc gtgtcgagca tcttcatctg ctccatcacg ctgtaaaaca catttgcacc
     8101 gcgagtctgc ccgtcctcca cgggttcaaa aacgtgaatg aacgaggcgc gctcactggc
     8161 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc
     8221 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
     8281 ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
     8341 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     8401 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     8461 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     8521 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     8581 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     8641 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     8701 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     8761 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     8821 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     8881 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     8941 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     9001 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     9061 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     9121 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     9181 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     9241 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     9301 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     9361 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     9421 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     9481 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     9541 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     9601 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     9661 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     9721 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     9781 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     9841 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     9901 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     9961 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
    10021 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
    10081 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
    10141 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
    10201 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
    10261 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
    10321 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
    10381 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
    10441 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
    10501 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
    10561 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
    10621 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
    10681 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
    10741 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
    10801 gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg
    10861 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac
    10921 catgattacg ccaagcgcgc ccgccgggta actcacgggg tatccatgtc catttctgcg
    10981 gcatccagcc aggatacccg tcctcgctga cgtaatatcc cagcgccgca ccgctgtcat
    11041 taatctgcac accggcacgg cagttccggc tgtcgccggt attgttcggg ttgctgatgc
    11101 gcttcgggct gaccatccgg aactgtgtcc ggaaaagccg cgacgaactg gtatcccagg
    11161 tggcctgaac gaacagttca ccgttaaagg cgtgcatggc cacaccttcc cgaatcatca
    11221 tggtaaacgt gcgttttcgc tcaacgtcaa tgcagcagca gtcatcctcg gcaaactctt
    11281 tccatgccgc ttcaacctcg cgggaaaagg cacgggcttc ttcctccccg atgcccagat
    11341 agcgccagct tgggcgatga ctgagccgga aaaaagaccc gacgatatga tcctgatgca
    11401 gctaga
//
