LOCUS       sequence-373450-       14306 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses FLAG tagged AsdCas12a VPR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14306)
  AUTHORS   Escobar M, Li J, Patel A, Liu S, Xu Q, Hilton IB
  TITLE     Quantification of Genome Editing and Transcriptional Control 
            Capabilities Reveals Hierarchies among Diverse CRISPR/Cas Systems in
            Human Cells.
  JOURNAL   ACS Synth Biol. 2022 Oct 21;11(10):3239-3250. doi: 
            10.1021/acssynbio.2c00156. Epub 2022 Sep 26.
  PUBMED    36162812
REFERENCE   2  (bases 1 to 14306)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acssynbio.2c00156"; journalName: "ACS Synth Biol"; date: 
            "2022-10-21- 21"; volume: "11"; issue: "10"; pages: "3239-3250"
COMMENT     Sequence Label: AsdCas12a VPR
FEATURES             Location/Qualifiers
     source          1..14306
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..199
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     154..174
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             217..397
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    444..569
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1062..1295
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1480..1524
                     /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    1822..1939
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_RNA        2071..2146
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2208..2419
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2444..6364
                     /codon_start=1
                     /product="CRISPR-associated protein from Acidaminococcus 
                     sp. BV3L6 (Zetsche et al., 2015)"
                     /label=AsCpf1
                     /translation="MTQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARND
                     HYKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRN
                     AIHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFD
                     KFTTYFSGFYENRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHF
                     ENVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVL
                     NLAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLL
                     RNENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTG
                     KITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTL
                     KKQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKA
                     RNYATKKPYSVEKFKLNFQMPTLASGWDVNKEKNNGAILFVKNGLYYLGIMPKQKGRYK
                     ALSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEP
                     LEITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSID
                     LSSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKG
                     HHGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLK
                     DQKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFF
                     HVPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIARGERNLIYITVIDSTGKILEQR
                     SLNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAV
                     VVLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQ
                     FTSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYD
                     VKTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIEN
                     HRFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQM
                     RNSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKES
                     KDLKLQNGISNQDWLAYIQELRN"
     regulatory      6084..6093
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             6365..6412
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6413..6436
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    6479..6503
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             6533..6682
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             6707..6727
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7157..7513
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALL"
     CDS             7532..8101
                     /codon_start=1
                     /product="transcriptional activation domain from the human 
                     herpesvirus 4 (Epstein-Barr virus) replication and 
                     transcription activator Rta/BRLF1 (Hardwick et al., 1992; 
                     Chavez et al., 2015)"
                     /label=Rta AD
                     /translation="RDSREGMFLPKPEAGSAISDVFEGREVCQPKRIRPFHPPGSPWAN
                     RPLPASLAPTPTGPVHEPVGSLTPAPVPQPLDPAPAVTPEASHLLEDPDEETSQAVKAL
                     REMADTVIPQKEEAAICGQMDLSHPPPRGHLDELTTTLESMTEDLNLDSPLTPELNEIL
                     DTFLNDECLLHAMHISTGLSIFDTSLF"
     CDS             8111..8167
                     /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             8168..8764
                     /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     8661..8681
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8780..9368
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8833..8853)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9251..9262)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9440..9673
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9699..9716)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9705..9929
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9975..10403
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10062..10081)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10272..10293
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10412..10432)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10417..10746
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10597..10732
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10659..10678
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10794..10841
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10860..11234
                     /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    11364..11485
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11401..11420)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11455..11474
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11534..11550)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11534..11550)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11547..11569)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11558..11574
                     /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(11582..11588,11589..11606,11607..11612))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11627..11648
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11765..11782)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11936..12524)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12016..12035)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12695..13486,13487..13555))
                     /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     13318..13337
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13556..13660)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(13735..13754)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        13926..14305
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
       61 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
      121 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
      181 gggaggtcta tataagcagc gcgttttgcc tgtactgggt ctctctggtt agaccagatc
      241 tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg
      301 ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc
      361 ctcagaccct tttagtcagt gtggaaaatc tctagcagtg gcgcccgaac agggacttga
      421 aagcgaaagg gaaaccagag gagctctctc gacgcaggac tcggcttgct gaagcgcgca
      481 cggcaagagg cgaggggcgg cgactggtga gtacgccaaa aattttgact agcggaggct
      541 agaaggagag agatgggtgc gagagcgtca gtattaagcg ggggagaatt agatcgcgat
      601 gggaaaaaat tcggttaagg ccagggggaa agaaaaaata taaattaaaa catatagtat
      661 gggcaagcag ggagctagaa cgattcgcag ttaatcctgg cctgttagaa acatcagaag
      721 gctgtagaca aatactggga cagctacaac catcccttca gacaggatca gaagaactta
      781 gatcattata taatacagta gcaaccctct attgtgtgca tcaaaggata gagataaaag
      841 acaccaagga agctttagac aagatagagg aagagcaaaa caaaagtaag accaccgcac
      901 agcaagcggc cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt
      961 gaattatata aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca
     1021 aagagaagag tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg
     1081 ttcttgggag cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc
     1141 agacaattat tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg
     1201 caacagcatc tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg
     1261 gctgtggaaa gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa
     1321 ctcatttgca ccactgctgt gccttggaat gctagttgga gtaataaatc tctggaacag
     1381 atttggaatc acacgacctg gatggagtgg gacagagaaa ttaacaatta cacaagctta
     1441 atacactcct taattgaaga atcgcaaaac cagcaagaaa agaatgaaca agaattattg
     1501 gaattagata aatgggcaag tttgtggaat tggtttaaca taacaaattg gctgtggtat
     1561 ataaaattat tcataatgat agtaggaggc ttggtaggtt taagaatagt ttttgctgta
     1621 ctttctatag tgaatagagt taggcaggga tattcaccat tatcgtttca gacccacctc
     1681 ccaaccccga ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac
     1741 agagacagat ccattcgatt agtgaacgga tcggcactgc gtgcgccaat tctgcagaca
     1801 aatggcagta ttcatccaca attttaaaag aaaagggggg attggggggt acagtgcagg
     1861 ggaaagaata gtagacataa tagcaacaga catacaaact aaagaattac aaaaacaaat
     1921 tacaaaaatt caaaattttc gggtttatta cagggacagc agagatccag tttggttaat
     1981 taaggtaccg atatcactag tgttccatta actgcgctaa aaataatttt taaatctttt
     2041 ttagcttctt gctctttttt gtacgtctct gttttagagc tagaaatagc aagttaaaat
     2101 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgaattcgc
     2161 tagctaggtc ttgaaaggag tgggaattgg ctccggtgcc cgtcagtggg cagagcgcac
     2221 atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgatccg gtgcctagag
     2281 aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga
     2341 gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg
     2401 gtttgccgcc agaacacagg accggttcta gagcgctgcc accatgacac agttcgaggg
     2461 ctttaccaac ctgtatcagg tgagcaagac actgcggttt gagctgatcc cacagggcaa
     2521 gaccctgaag cacatccagg agcagggctt catcgaggag gacaaggccc gcaatgatca
     2581 ctacaaggag ctgaagccca tcatcgatcg gatctacaag acctatgccg accagtgcct
     2641 gcagctggtg cagctggatt gggagaacct gagcgccgcc atcgactcct atagaaagga
     2701 gaaaaccgag gagacaagga acgccctgat cgaggagcag gccacatatc gcaatgccat
     2761 ccacgactac ttcatcggcc ggacagacaa cctgaccgat gccatcaata agagacacgc
     2821 cgagatctac aagggcctgt tcaaggccga gctgtttaat ggcaaggtgc tgaagcagct
     2881 gggcaccgtg accacaaccg agcacgagaa cgccctgctg cggagcttcg acaagtttac
     2941 aacctacttc tccggctttt atgagaacag gaagaacgtg ttcagcgccg aggatatcag
     3001 cacagccatc ccacaccgca tcgtgcagga caacttcccc aagtttaagg agaattgtca
     3061 catcttcaca cgcctgatca ccgccgtgcc cagcctgcgg gagcactttg agaacgtgaa
     3121 gaaggccatc ggcatcttcg tgagcacctc catcgaggag gtgttttcct tcccttttta
     3181 taaccagctg ctgacacaga cccagatcga cctgtataac cagctgctgg gaggaatctc
     3241 tcgggaggca ggcaccgaga agatcaaggg cctgaacgag gtgctgaatc tggccatcca
     3301 gaagaatgat gagacagccc acatcatcgc ctccctgcca cacagattca tccccctgtt
     3361 taagcagatc ctgtccgata ggaacaccct gtctttcatc ctggaggagt ttaagagcga
     3421 cgaggaagtg atccagtcct tctgcaagta caagacactg ctgagaaacg agaacgtgct
     3481 ggagacagcc gaggccctgt ttaacgagct gaacagcatc gacctgacac acatcttcat
     3541 cagccacaag aagctggaga caatcagcag cgccctgtgc gaccactggg atacactgag
     3601 gaatgccctg tatgagcgga gaatctccga gctgacaggc aagatcacca agtctgccaa
     3661 ggagaaggtg cagcgcagcc tgaagcacga ggatatcaac ctgcaggaga tcatctctgc
     3721 cgcaggcaag gagctgagcg aggccttcaa gcagaaaacc agcgagatcc tgtcccacgc
     3781 acacgccgcc ctggatcagc cactgcctac aaccctgaag aagcaggagg agaaggagat
     3841 cctgaagtct cagctggaca gcctgctggg cctgtaccac ctgctggact ggtttgccgt
     3901 ggatgagtcc aacgaggtgg accccgagtt ctctgcccgg ctgaccggca tcaagctgga
     3961 gatggagcct tctctgagct tctacaacaa ggccagaaat tatgccacca agaagcccta
     4021 ctccgtggag aagttcaagc tgaactttca gatgcctaca ctggcctctg gctgggacgt
     4081 gaataaggag aagaacaatg gcgccatcct gtttgtgaag aacggcctgt actatctggg
     4141 catcatgcca aagcagaagg gcaggtataa ggccctgagc ttcgagccca cagagaaaac
     4201 cagcgagggc tttgataaga tgtactatga ctacttccct gatgccgcca agatgatccc
     4261 aaagtgcagc acccagctga aggccgtgac agcccacttt cagacccaca caacccccat
     4321 cctgctgtcc aacaatttca tcgagcctct ggagatcaca aaggagatct acgacctgaa
     4381 caatcctgag aaggagccaa agaagtttca gacagcctac gccaagaaaa ccggcgacca
     4441 gaagggctac agagaggccc tgtgcaagtg gatcgacttc acaagggatt ttctgtccaa
     4501 gtataccaag acaacctcta tcgatctgtc tagcctgcgg ccatcctctc agtataagga
     4561 cctgggcgag tactatgccg agctgaatcc cctgctgtac cacatcagct tccagagaat
     4621 cgccgagaag gagatcatgg atgccgtgga gacaggcaag ctgtacctgt tccagatcta
     4681 taacaaggac tttgccaagg gccaccacgg caagcctaat ctgcacacac tgtattggac
     4741 cggcctgttt tctccagaga acctggccaa gacaagcatc aagctgaatg gccaggccga
     4801 gctgttctac cgccctaagt ccaggatgaa gaggatggca caccggctgg gagagaagat
     4861 gctgaacaag aagctgaagg atcagaaaac cccaatcccc gacaccctgt accaggagct
     4921 gtacgactat gtgaatcaca gactgtccca cgacctgtct gatgaggcca gggccctgct
     4981 gcccaacgtg atcaccaagg aggtgtctca cgagatcatc aaggataggc gctttaccag
     5041 cgacaagttc tttttccacg tgcctatcac actgaactat caggccgcca attccccatc
     5101 taagttcaac cagagggtga atgcctacct gaaggagcac cccgagacac ctatcatcgg
     5161 catcgcccgg ggcgagagaa acctgatcta tatcacagtg atcgactcca ccggcaagat
     5221 cctggagcag cggagcctga acaccatcca gcagtttgat taccagaaga agctggacaa
     5281 cagggagaag gagagggtgg cagcaaggca ggcctggtct gtggtgggca caatcaagga
     5341 tctgaagcag ggctatctga gccaggtcat ccacgagatc gtggacctga tgatccacta
     5401 ccaggccgtg gtggtgctgg agaacctgaa tttcggcttt aagagcaaga ggaccggcat
     5461 cgccgagaag gccgtgtacc agcagttcga gaagatgctg atcgataagc tgaattgcct
     5521 ggtgctgaag gactatccag cagagaaagt gggaggcgtg ctgaacccat accagctgac
     5581 agaccagttc acctcctttg ccaagatggg cacccagtct ggcttcctgt tttacgtgcc
     5641 tgccccatat acatctaaga tcgatcccct gaccggcttc gtggacccct tcgtgtggaa
     5701 aaccatcaag aatcacgaga gccgcaagca cttcctggag ggcttcgact ttctgcacta
     5761 cgacgtgaaa accggcgact tcatcctgca ctttaagatg aacagaaatc tgtccttcca
     5821 gaggggcctg cccggcttta tgcctgcatg ggatatcgtg ttcgagaaga acgagacaca
     5881 gtttgacgcc aagggcaccc ctttcatcgc cggcaagaga atcgtgccag tgatcgagaa
     5941 tcacagattc accggcagat accgggacct gtatcctgcc aacgagctga tcgccctgct
     6001 ggaggagaag ggcatcgtgt tcagggatgg ctccaacatc ctgccaaagc tgctggagaa
     6061 tgacgattct cacgccatcg acaccatggt ggccctgatc cgcagcgtgc tgcagatgcg
     6121 gaactccaat gccgccacag gcgaggacta tatcaacagc cccgtgcgcg atctgaatgg
     6181 cgtgtgcttc gactcccggt ttcagaaccc agagtggccc atggacgccg atgccaatgg
     6241 cgcctaccac atcgccctga agggccagct gctgctgaat cacctgaagg agagcaagga
     6301 tctgaagctg cagaacggca tctccaatca ggactggctg gcctacatcc aggagctgcg
     6361 caacaagcga cctgccgcca caaagaaggc tggacaggct aagaagaaga aagattacaa
     6421 agacgatgac gataagggat cctcgccagg gatccgtcga cttgacgcgt tgatatcaac
     6481 aagtttgtac aaaaaagcag gctacaaaga ggccagcggt tccggacggg ctgacgcatt
     6541 ggacgatttt gatctggata tgctgggaag tgacgccctc gatgattttg accttgacat
     6601 gcttggttcg gatgcccttg atgactttga cctcgacatg ctcggcagtg acgcccttga
     6661 tgatttcgac ctggacatgc tgattaactc tagaagttcc ggatctccga aaaagaaacg
     6721 caaagttggt agccagtacc tgcccgacac cgacgaccgg caccggatcg aggaaaagcg
     6781 gaagcggacc tacgagacat tcaagagcat catgaagaag tcccccttca gcggccccac
     6841 cgaccctaga cctccaccta gaagaatcgc cgtgcccagc agatccagcg ccagcgtgcc
     6901 aaaacctgcc ccccagcctt accccttcac cagcagcctg agcaccatca actacgacga
     6961 gttccctacc atggtgttcc ccagcggcca gatctctcag gcctctgctc tggctccagc
     7021 ccctcctcag gtgctgcctc aggctcctgc tcctgcacca gctccagcca tggtgtctgc
     7081 actggctcag gcaccagcac ccgtgcctgt gctggctcct ggacctccac aggctgtggc
     7141 tccaccagcc cctaaaccta cacaggccgg cgagggcaca ctgtctgaag ctctgctgca
     7201 gctgcagttc gacgacgagg atctgggagc cctgctggga aacagcaccg atcctgccgt
     7261 gttcaccgac ctggccagcg tggacaacag cgagttccag cagctgctga accagggcat
     7321 ccctgtggcc cctcacacca ccgagcccat gctgatggaa taccccgagg ccatcacccg
     7381 gctcgtgaca ggcgctcaga ggcctcctga tccagctcct gcccctctgg gagcaccagg
     7441 cctgcctaat ggactgctgt ctggcgacga ggacttcagc tctatcgccg atatggattt
     7501 ctcagccttg ctgggctctg gcagcggcag ccgggattcc agggaaggga tgtttttgcc
     7561 gaagcctgag gccggctccg ctattagtga cgtgtttgag ggccgcgagg tgtgccagcc
     7621 aaaacgaatc cggccatttc atcctccagg aagtccatgg gccaaccgcc cactccccgc
     7681 cagcctcgca ccaacaccaa ccggtccagt acatgagcca gtcgggtcac tgaccccggc
     7741 accagtccct cagccactgg atccagcgcc cgcagtgact cccgaggcca gtcacctgtt
     7801 ggaggatccc gatgaagaga cgagccaggc tgtcaaagcc cttcgggaga tggccgatac
     7861 tgtgattccc cagaaggaag aggctgcaat ctgtggccaa atggaccttt cccatccgcc
     7921 cccaaggggc catctggatg agctgacaac cacacttgag tccatgaccg aggatctgaa
     7981 cctggactca cccctgaccc cggaattgaa cgagattctg gataccttcc tgaacgacga
     8041 gtgcctcttg catgccatgc atatcagcac aggactgtcc atcttcgaca catctctgtt
     8101 tggatccggc gcaacaaact tctctctgct gaaacaagcc ggagatgtcg aagagaatcc
     8161 tggaccgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc
     8221 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc
     8281 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
     8341 cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc
     8401 ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag
     8461 cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa
     8521 ggagcccgcg tggttcctgg ccaccgtcgg agtctcgccc gaccaccagg gcaagggtct
     8581 gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt
     8641 cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac
     8701 cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc
     8761 ctgaacgcgt taagtcgaca atcaacctct ggattacaaa atttgtgaaa gattgactgg
     8821 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     8881 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     8941 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     9001 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     9061 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     9121 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc
     9181 gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
     9241 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     9301 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     9361 ctccccgcgt cgactttaag accaatgact tacaaggcag ctgtagatct tagccacttt
     9421 ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca agatctgctt
     9481 tttgcttgta ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa
     9541 ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt
     9601 gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg
     9661 aaaatctcta gcagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg
     9721 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     9781 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     9841 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     9901 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     9961 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
    10021 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
    10081 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
    10141 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
    10201 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
    10261 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
    10321 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
    10381 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
    10441 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
    10501 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
    10561 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
    10621 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
    10681 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
    10741 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcagca cgtgttgaca
    10801 attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg aactaaacca
    10861 tggccaagtt gaccagtgcc gttccggtgc tcaccgcgcg cgacgtcgcc ggagcggtcg
    10921 agttctggac cgaccggctc gggttctccc gggacttcgt ggaggacgac ttcgccggtg
    10981 tggtccggga cgacgtgacc ctgttcatca gcgcggtcca ggaccaggtg gtgccggaca
    11041 acaccctggc ctgggtgtgg gtgcgcggcc tggacgagct gtacgccgag tggtcggagg
    11101 tcgtgtccac gaacttccgg gacgcctccg ggccggccat gaccgagatc ggcgagcagc
    11161 cgtgggggcg ggagttcgcc ctgcgcgacc cggccggcaa ctgcgtgcac ttcgtggccg
    11221 aggagcagga ctgacacgtg ctacgagatt tcgattccac cgccgccttc tatgaaaggt
    11281 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
    11341 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
    11401 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
    11461 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
    11521 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
    11581 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
    11641 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
    11701 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
    11761 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
    11821 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
    11881 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
    11941 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
    12001 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
    12061 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
    12121 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
    12181 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
    12241 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
    12301 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
    12361 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
    12421 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
    12481 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
    12541 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
    12601 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
    12661 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
    12721 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
    12781 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
    12841 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
    12901 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
    12961 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
    13021 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
    13081 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
    13141 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
    13201 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
    13261 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
    13321 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
    13381 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
    13441 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
    13501 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
    13561 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
    13621 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
    13681 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
    13741 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
    13801 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
    13861 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
    13921 gcgttgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
    13981 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
    14041 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
    14101 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
    14161 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
    14221 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
    14281 cgtattagtc atcgctatta ccatgg
//
