LOCUS       sequence-231716-       13973 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  U6-sgRNA EF1a-ProCas9-Flavi-P2A-Puro.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13973)
  AUTHORS   Oakes BL, Fellmann C, Rishi H, Taylor KL, Ren SM, Nadler DC, Yokoo 
            R, Arkin AP, Doudna JA, Savage DF
  TITLE     CRISPR-Cas9 Circular Permutants as Programmable Scaffolds for Genome
            Modification.
  JOURNAL   Cell. 2019 Jan 10;176(1-2):254-267.e16. doi: 
            10.1016/j.cell.2018.11.052.
  PUBMED    30633905
REFERENCE   2  (bases 1 to 13973)
  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: "Cell."; 
            date: "2019-01-10"; volume: "176(1-2):254-267.e16. doi"; pages: " 
            10.1016/j.cell.2018.11.052"
COMMENT     Sequence Label: pCF712
FEATURES             Location/Qualifiers
     source          1..13973
                     /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        2608..2856
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2608..2628
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2779..2798
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        3012..4189
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3243..4180
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4137..4157
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      4196..4205
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4208..4228
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5246..7744
                     /codon_start=1
                     /product="C-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(C)
                     /translation="KPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEI
                     SGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTY
                     AHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIH
                     DDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPE
                     NIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYL
                     QNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSEEV
                     VKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQI
                     LDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVV
                     GTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITL
                     ANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESIL
                     PKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIME
                     RSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALP
                     SKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLD
                     KVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATL
                     IHQSITGLYETRIDLSQLGGD"
     CDS             8366..8386
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8396..8452
                     /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             8453..9049
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     8946..8966
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    9065..9653
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9118..9138)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9536..9547)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9725..9958
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9984..10001)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9990..10214
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10260..10688
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10347..10366)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10557..10578
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10697..10717)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10702..11031
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10882..11017
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10944..10963
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11079..11126
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11145..11519
                     /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    11649..11770
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11686..11705)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11740..11759
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11819..11835)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11819..11835)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11832..11854)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11843..11859
                     /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(11867..11873,11874..11891,11892..11897))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11912..11933
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12050..12067)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12221..12809)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12301..12320)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12980..13771,13772..13840))
                     /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     13603..13622
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13841..13945)
                     /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 aggtaccgag ggcctatttc ccatgattcc ttcatatttg
     2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag
     2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
     2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
     2821 ttggctttat atatcttgtg gaaaggacga aacaccggag acggaggacg acgaacgtct
     2881 ctgtttaaga gctatgctgg aaacagcata gcaagtttaa ataaggctag tccgttatca
     2941 acttgaaaaa gtggcaccga gtcggtgctt tttttgaatt cgctagctag gtcttgaaag
     3001 gagtgggaat tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg
     3061 agaagttggg gggaggggtc ggcaattgat ccggtgccta gagaaggtgg cgcggggtaa
     3121 actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt
     3181 atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac
     3241 aggtaagtgc cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg
     3301 tgccttgaat tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg
     3361 aagtgggtgg gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt
     3421 tgaggcctgg cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg
     3481 tctcgctgct ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct
     3541 ttttttctgg caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt
     3601 ttttggggcc gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg
     3661 gggcctgcga gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc
     3721 tctggtgcct ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg
     3781 gtcggcacca gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc
     3841 aaaatggagg acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag
     3901 ggcctttccg tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag
     3961 gcacctcgat tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt
     4021 ttatgcgatg gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca
     4081 cttgatgtaa ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa
     4141 gcctcagaca gtggttcaaa gtttttttct tccatttcag gtgtcgtgaa ccggtgccac
     4201 catggcccca aagaagaagc ggaaggtcag cgctaacccc atcaacgcca gcggcgtgga
     4261 cgccaaggcc atcctgtctg ccagactgag caagagcaga cggctggaaa atctgatcgc
     4321 ccagctgccc ggcgagaaga agaatggcct gttcggaaac ctgattgccc tgagcctggg
     4381 cctgaccccc aacttcaaga gcaacttcga cctggccgag gatgccaaac tgcagctgag
     4441 caaggacacc tacgacgacg acctggacaa cctgctggcc cagatcggcg accagtacgc
     4501 cgacctgttt ctggccgcca agaacctgtc cgacgccatc ctgctgagcg acatcctgag
     4561 agtgaacacc gagatcacca aggcccccct gagcgcctct atgatcaaga gatacgacga
     4621 gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta
     4681 caaagagatt ttcttcgacc agagcaagaa cggctacgcc ggctacattg acggcggagc
     4741 cagccaggaa gagttctaca agttcatcaa gcccatcctg gaaaagatgg acggcaccga
     4801 ggaactgctc gtgaagctga acagagagga cctgctgcgg aagcagcgga ccttcgacaa
     4861 cggcagcatc ccccaccaga tccacctggg agagctgcac gccattctgc ggcggcagga
     4921 agatttttac ccattcctga aggacaaccg ggaaaagatc gagaagatcc tgaccttccg
     4981 catcccctac tacgtgggcc ctctggccag gggaaacagc agattcgcct ggatgaccag
     5041 aaagagcgag gaaaccatca ccccctggaa cttcgaggaa gtggtggaca agggcgcttc
     5101 cgcccagagc ttcatcgagc ggatgaccaa cttcgataag aacctgccca acgagaaggt
     5161 gctgcccaag cacagcctgc tgtacgagta cttcaccgtg tataacgagc tgaccaaagt
     5221 gaaatacgtg accgagggaa tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc
     5281 catcgtggac ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga
     5341 ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt
     5401 caacgcctcc ctgggcacat accacgatct gctgaaaatt atcaaggaca aggacttcct
     5461 ggacaatgag gaaaacgagg acattctgga agatatcgtg ctgaccctga cactgtttga
     5521 ggacagagag atgatcgagg aacggctgaa aacctatgcc cacctgttcg acgacaaagt
     5581 gatgaagcag ctgaagcggc ggagatacac cggctggggc aggctgagcc ggaagctgat
     5641 caacggcatc cgggacaagc agtccggcaa gacaatcctg gatttcctga agtccgacgg
     5701 cttcgccaac agaaacttca tgcagctgat ccacgacgac agcctgacct ttaaagagga
     5761 catccagaaa gcccaggtgt ccggccaggg cgatagcctg cacgagcaca ttgccaatct
     5821 ggccggcagc cccgccatta agaagggcat cctgcagaca gtgaaggtgg tggacgagct
     5881 cgtgaaagtg atgggccggc acaagcccga gaacatcgtg atcgaaatgg ccagagagaa
     5941 ccagaccacc cagaagggac agaagaacag ccgcgagaga atgaagcgga tcgaagaggg
     6001 catcaaagag ctgggcagcc agatcctgaa agaacacccc gtggaaaaca cccagctgca
     6061 gaacgagaag ctgtacctgt actacctgca gaatgggcgg gatatgtacg tggaccagga
     6121 actggacatc aaccggctgt ccgactacga tgtggaccat atcgtgcctc agagctttct
     6181 gaaggacgac tccatcgaca acaaggtgct gaccagaagc gacaagaacc ggggcaagag
     6241 cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag aactactggc ggcagctgct
     6301 gaacgccaag ctgattaccc agagaaagtt cgacaatctg accaaggccg agagaggcgg
     6361 cctgagcgaa ctggataagg ccggcttcat caagagacag ctggtggaaa cccggcagat
     6421 cacaaagcac gtggcacaga tcctggactc ccggatgaac actaagtacg acgagaatga
     6481 caagctgatc cgggaagtga aagtgatcac cctgaagtcc aagctggtgt ccgatttccg
     6541 gaaggatttc cagttttaca aagtgcgcga gatcaacaac taccaccacg cccacgacgc
     6601 ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc tggaaagcga
     6661 gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag atgatcgcca agagcgagca
     6721 ggaaatcggc aaggctaccg ccaagtactt cttctacagc aacatcatga actttttcaa
     6781 gaccgagatt accctggcca acggcgagat ccggaagcgg cctctgatcg agacaaacgg
     6841 cgaaaccggg gagatcgtgt gggataaggg ccgggatttt gccaccgtgc ggaaagtgct
     6901 gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa
     6961 agagtctatc ctgcccaaga ggaacagcga taagctgatc gccagaaaga aggactggga
     7021 ccctaagaag tacggcggct tcgacagccc caccgtggcc tattctgtgc tggtggtggc
     7081 caaagtggaa aagggcaagt ccaagaaact gaagagtgtg aaagagctgc tggggatcac
     7141 catcatggaa agaagcagct tcgagaagaa tcccatcgac tttctggaag ccaagggcta
     7201 caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag tactccctgt tcgagctgga
     7261 aaacggccgg aagagaatgc tggcctctgc cggcgaactg cagaagggaa acgaactggc
     7321 cctgccctcc aaatatgtga acttcctgta cctggccagc cactatgaga agctgaaggg
     7381 ctcccccgag gataatgagc agaaacagct gtttgtggaa cagcacaagc actacctgga
     7441 cgagatcatc gagcagatca gcgagttctc caagagagtg atcctggccg acgctaatct
     7501 ggacaaagtg ctgtccgcct acaacaagca ccgggataag cccatcagag agcaggccga
     7561 gaatatcatc cacctgttta ccctgaccaa tctgggagcc cctgccgcct tcaagtactt
     7621 tgacaccacc atcgaccgga agaggtacac cagcaccaaa gaggtgctgg acgccaccct
     7681 gatccaccag agcatcaccg gcctgtacga gacacggatc gacctgtctc agctgggagg
     7741 cgacaaacaa aagaaacgcg gaggcaagga caagaagtac agcatcggcc tggacatcgg
     7801 caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca gcaagaaatt
     7861 caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg gagccctgct
     7921 gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca gaagaagata
     7981 caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg agatggccaa
     8041 ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag aggataagaa
     8101 gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc acgagaagta
     8161 ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg ccgacctgcg
     8221 gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc tgatcgaggg
     8281 cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg tgcagaccta
     8341 caaccagctg ttcgaggaaa ctagtccaaa aaagaagaga aaggtaggat ccggcgcaac
     8401 aaacttctct ctgctgaaac aagccggaga tgtcgaagag aatcctggac cgaccgagta
     8461 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc
     8521 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga
     8581 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
     8641 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
     8701 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
     8761 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
     8821 cctggccacc gtcggagtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
     8881 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc
     8941 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
     9001 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgaa cgcgttaagt
     9061 cgacaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     9121 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     9181 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     9241 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     9301 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     9361 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     9421 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     9481 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     9541 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     9601 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcgtcgact
     9661 ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg
     9721 ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc ttgtactggg
     9781 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
     9841 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
     9901 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagg
     9961 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
    10021 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
    10081 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
    10141 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
    10201 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
    10261 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
    10321 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
    10381 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
    10441 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
    10501 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
    10561 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
    10621 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
    10681 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
    10741 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
    10801 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
    10861 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
    10921 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
    10981 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
    11041 ggagcttgta tatccatttt cggatctgat cagcacgtgt tgacaattaa tcatcggcat
    11101 agtatatcgg catagtataa tacgacaagg tgaggaacta aaccatggcc aagttgacca
    11161 gtgccgttcc ggtgctcacc gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc
    11221 ggctcgggtt ctcccgggac ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg
    11281 tgaccctgtt catcagcgcg gtccaggacc aggtggtgcc ggacaacacc ctggcctggg
    11341 tgtgggtgcg cggcctggac gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact
    11401 tccgggacgc ctccgggccg gccatgaccg agatcggcga gcagccgtgg gggcgggagt
    11461 tcgccctgcg cgacccggcc ggcaactgcg tgcacttcgt ggccgaggag caggactgac
    11521 acgtgctacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
    11581 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
    11641 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
    11701 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
    11761 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt
    11821 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
    11881 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
    11941 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
    12001 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
    12061 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
    12121 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
    12181 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
    12241 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
    12301 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
    12361 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
    12421 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
    12481 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
    12541 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
    12601 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
    12661 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
    12721 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
    12781 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
    12841 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
    12901 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
    12961 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
    13021 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
    13081 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
    13141 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
    13201 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
    13261 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
    13321 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
    13381 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
    13441 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
    13501 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
    13561 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
    13621 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
    13681 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
    13741 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
    13801 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
    13861 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
    13921 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gac
//
