LOCUS       sequence-426762-       14982 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of multiAsCas12a-KRAB.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14982)
  AUTHORS   Hsiung CC, Wilson CM, Sambold NA, Dai R, Chen Q, Teyssier N, 
            Misiukiewicz S, Arab A, O'Loughlin T, Cofsky JC, Shi J, Gilbert LA
  TITLE     Engineered CRISPR-Cas12a for higher-order combinatorial chromatin 
            perturbations.
  JOURNAL   Nat Biotechnol. 2024 May 17. doi: 10.1038/s41587-024-02224-0.
  PUBMED    38760567
REFERENCE   2  (bases 1 to 14982)
  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: "Nat 
            Biotechnol. 2024 May 17. doi: 10.1038/s41587-024-02224-0."
COMMENT     Sequence Label: pCH45 (multiAsCas12a-KRAB lenti)
FEATURES             Location/Qualifiers
     source          1..14982
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(23..41)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(24..41)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(331..350)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(564..583)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     683..705
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(743..761)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        829..933
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(934..1002,1003..1794)
                     /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(1152..1171)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1965..2553
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2454..2473
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2707..2724
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2794..2814)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2799..3128
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2979..3114
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          4262..4327
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             4457..4477
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    4902..5036
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4939..4958)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4993..5012
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             5204..5837
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    5884..6009
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6506..6739
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6924..6968
                     /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    7253..7370
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        8112..8519
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     CDS             8562..8588
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             8598..8624
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             8634..8660
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             8670..8696
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             8706..8732
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             8742..8768
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             8778..12695
                     /codon_start=1
                     /product="CRISPR-associated protein from Acidaminococcus 
                     sp. BV3L6 (Zetsche et al., 2015)"
                     /label=AsCpf1
                     /translation="TQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARNDH
                     YKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRNA
                     IHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFDK
                     FTTYFSGFYRNRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHFE
                     NVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVLN
                     LAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLLR
                     NENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTGK
                     ITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTLK
                     KQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKAR
                     NYATKKPYSVEKFKLNFQMPTLARGWDVNREKNNGAILFVKNGLYYLGIMPKQKGRYKA
                     LSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEPL
                     EITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSIDL
                     SSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKGH
                     HGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLKD
                     QKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFFH
                     VPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIDRGERNLIYITVIDSTGKILEQRS
                     LNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAVV
                     VLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQF
                     TSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYDV
                     KTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIENH
                     RFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQMA
                     NSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKESK
                     DLKLQNGISNQDWLAYIQELRN"
     CDS             12708..12734
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             12744..12764
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             13041..13226
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             13236..13292
                     /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             13293..14000
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDM
                     ALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVA
                     RYCDLPSKLGHKLN"
     misc_feature    14049..14637
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(14102..14122)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(14520..14531)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(14640..14656)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(14641..14657)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             14747..14980
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
ORIGIN
        1 atctagaatt aattccgtgt attctatagt gtcacctaaa tcgtatgtgt atgatacata
       61 aggttatgta ttaattgtag ccgcgttcta acgacaatat gtacaagcct aattgtgtag
      121 catctggctt actgaagcag accctatcat ctctctcgta aactgccgtc agagtcggtt
      181 tggttggacg aaccttctga gtttctggta acgccgtccc gcacccggaa atggtcagcg
      241 aaccaatcag cagggtcatc gctagccaga tcctctacgc cggacgcatc gtggccggca
      301 tcaccggcgc cacaggtgcg gttgctggcg cctatatcgc cgacatcacc gatggggaag
      361 atcgggctcg ccacttcggg ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc
      421 ccgtggccgg gggactgttg ggcgccatct ccttgcatgc accattcctt gcggcggcgg
      481 tgctcaacgg cctcaaccta ctactgggct gcttcctaat gcaggagtcg cataagggag
      541 agcgtcgaat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
      601 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
      661 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
      721 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
      781 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
      841 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
      901 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
      961 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     1021 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     1081 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     1141 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     1201 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     1261 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     1321 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     1381 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     1441 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     1501 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     1561 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     1621 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     1681 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     1741 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     1801 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     1861 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     1921 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     1981 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     2041 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     2101 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     2161 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     2221 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     2281 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     2341 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     2401 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     2461 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     2521 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     2581 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     2641 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     2701 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     2761 cccgcgcgtt ggccgattca ttaatgcagc tgtggaatgt gtgtcagtta gggtgtggaa
     2821 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2881 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     2941 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3001 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3061 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3121 tttgcaaaaa gcttggacac aagacaggct tgcgagatat gtttgagaat accactttat
     3181 cccgcgtcag ggagaggcag tgcgtaaaaa gacgcggact catgtgaaat actggttttt
     3241 agtgcgccag atctctataa tctcgcgcaa cctattttcc cctcgaacac tttttaagcc
     3301 gtagataaac aggctgggac acttcacatg agcgaaaaat acatcgtcac ctgggacatg
     3361 ttgcagatcc atgcacgtaa actcgcaagc cgactgatgc cttctgaaca atggaaaggc
     3421 attattgccg taagccgtgg cggtctgtac cgggtgcgtt actggcgcgt gaactgggta
     3481 ttcgtcatgt cgataccgtt tgtatttcca gctacgatca cgacaaccag cgcgagctta
     3541 aagtgctgaa acgcgcagaa ggcgatggcg aaggcttcat cgttattgat gacctggtgg
     3601 ataccggtgg tactgcggtt gcgattcgtg aaatgtatcc aaaagcgcac tttgtcacca
     3661 tcttcgcaaa accggctggt cgtccgctgg ttgatgacta tgttgttgat atcccgcaag
     3721 atacctggat tgaacagccg tgggatatgg gcgtcgtatt cgtcccgcca atctccggtc
     3781 gctaatcttt tcaacgcctg gcactgccgg gcgttgttct ttttaacttc aggcgggtta
     3841 caatagtttc cagtaagtat tctggaggct gcatccatga cacaggcaaa cctgagcgaa
     3901 accctgttca aaccccgctt taaacatcct gaaacctcga cgctagtccg ccgctttaat
     3961 cacggcgcac aaccgcctgt gcagtcggcc cttgatggta aaaccatccc tcactggtat
     4021 cgcatgatta accgtctgat gtggatctgg cgcggcattg acccacgcga aatcctcgac
     4081 gtccaggcac gtattgtgat gagcgatgcc gaacgtaccg acgatgattt atacgatacg
     4141 gtgattggct accgtggcgg caactggatt tatgagtggg ccccggatct ttgtgaagga
     4201 accttacttc tgtggtgtga cataattgga caaactacct acagagattt aaagctctaa
     4261 ggtaaatata aaatttttaa gtgtataatg tgttaaacta ctgattctaa ttgtttgtgt
     4321 attttagatt ccaacctatg gaactgatga atgggagcag tggtggaatg cctttaatga
     4381 ggaaaacctg ttttgctcag aagaaatgcc atctagtgat gatgaggcta ctgctgactc
     4441 tcaacattct actcctccaa aaaagaagag aaaggtagaa gaccccaagg actttccttc
     4501 agaattgcta agttttttga gtcatgctgt gtttagtaat agaactcttg cttgctttgc
     4561 tatttacacc acaaaggaaa aagctgcact gctatacaag aaaattatgg aaaaatattc
     4621 tgtaaccttt ataagtaggc ataacagtta taatcataac atactgtttt ttcttactcc
     4681 acacaggcat agagtgtctg ctattaataa ctatgctcaa aaattgtgta cctttagctt
     4741 tttaatttgt aaaggggtta ataaggaata tttgatgtat agtgccttga ctagagatca
     4801 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     4861 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     4921 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     4981 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatcaact
     5041 ggataactca agctaaccaa aatcatccca aacttcccac cccataccct attaccactg
     5101 ccaattacct gtggtttcat ttactctaaa cctgtgattc ctctgaatta ttttcatttt
     5161 aaagaaattg tatttgttaa atatgtacta caaacttagt agttggaagg gctaattcac
     5221 tcccaaagaa gacaagatat ccttgatctg tggatctacc acacacaagg ctacttccct
     5281 gattagcaga actacacacc agggccaggg gtcagatatc cactgacctt tggatggtgc
     5341 tacaagctag taccagttga gccagataag gtagaagagg ccaataaagg agagaacacc
     5401 agcttgttac accctgtgag cctgcatggg atggatgacc cggagagaga agtgttagag
     5461 tggaggtttg acagccgcct agcatttcat cacgtggccc gagagctgca tccggagtac
     5521 ttcaagaact gctgatatcg agcttgctac aagggacttt ccgctgggga ctttccaggg
     5581 aggcgtggcc tgggcgggac tggggagtgg cgagccctca gatcctgcat ataagcagct
     5641 gctttttgcc tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg
     5701 ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt
     5761 gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt
     5821 gtggaaaatc tctagcagtg gcgcccgaac agggacttga aagcgaaagg gaaaccagag
     5881 gagctctctc gacgcaggac tcggcttgct gaagcgcgca cggcaagagg cgaggggcgg
     5941 cgactggtga gtacgccaaa aattttgact agcggaggct agaaggagag agatgggtgc
     6001 gagagcgtca gtattaagcg ggggagaatt agatcgcgat gggaaaaaat tcggttaagg
     6061 ccagggggaa agaaaaaata taaattaaaa catatagtat gggcaagcag ggagctagaa
     6121 cgattcgcag ttaatcctgg cctgttagaa acatcagaag gctgtagaca aatactggga
     6181 cagctacaac catcccttca gacaggatca gaagaactta gatcattata taatacagta
     6241 gcaaccctct attgtgtgca tcaaaggata gagataaaag acaccaagga agctttagac
     6301 aagatagagg aagagcaaaa caaaagtaag accaccgcac agcaagcggc cggccgctga
     6361 tcttcagacc tggaggagga gatatgaggg acaattggag aagtgaatta tataaatata
     6421 aagtagtaaa aattgaacca ttaggagtag cacccaccaa ggcaaagaga agagtggtgc
     6481 agagagaaaa aagagcagtg ggaataggag ctttgttcct tgggttcttg ggagcagcag
     6541 gaagcactat gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa ttattgtctg
     6601 gtatagtgca gcagcagaac aatttgctga gggctattga ggcgcaacag catctgttgc
     6661 aactcacagt ctggggcatc aagcagctcc aggcaagaat cctggctgtg gaaagatacc
     6721 taaaggatca acagctcctg gggatttggg gttgctctgg aaaactcatt tgcaccactg
     6781 ctgtgccttg gaatgctagt tggagtaata aatctctgga acagatttgg aatcacacga
     6841 cctggatgga gtgggacaga gaaattaaca attacacaag cttaatacac tccttaattg
     6901 aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt attggaatta gataaatggg
     6961 caagtttgtg gaattggttt aacataacaa attggctgtg gtatataaaa ttattcataa
     7021 tgatagtagg aggcttggta ggtttaagaa tagtttttgc tgtactttct atagtgaata
     7081 gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac
     7141 ccgacaggcc cgaaggaata gaagaagaag gtggagagag agacagagac agatccattc
     7201 gattagtgaa cggatctcga cggtatcgcc aaatggcagt attcatccac aattttaaaa
     7261 gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag
     7321 acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt
     7381 acagggacag cagagatcca gtttggatcg ataagcttga tatcgaattc gggaggtggt
     7441 ccctgcagtt acgccaatga taacccccgc cagaaaaatc ttagtagcct tccctttttg
     7501 ttttccgtgc cccaactcgg cggattgact cggccccttc cggaaacacc cgaatcaact
     7561 tctagtcaaa ttattgttca cgccgcaatg acccacccct ggcccgcgtc tgtggaactg
     7621 acccctggtg tacaggagag ttcgctgctg aaagtggtcc caaaggggta ctagttttta
     7681 agctcccaac tccccctccc ccagcgtctg gaggattcca caccctcgca ccgcaggggc
     7741 gaggaagtgg gcggagtccg gttttggcgc cagccgctga ggctgccaag cagaaaagcc
     7801 accgctgagg agactccggt cactgtcctc gccccgcctc ccccttccct ccccttgggg
     7861 accaccgggc gccacgccgc gaacggtaag tgccgcggtc gtcggcgcct ccgccctccc
     7921 cctagggccc caattcccag cgggcgcggc gcgcggcccc gcgcgcagct cccggctccc
     7981 tcccccttcg gatgtggctt gagctgtagg cgcggagtcc tgcagccccg ataaaataaa
     8041 agattttatt tagtctccag aaaaaggggg gaatgaaaga ccccacctgt aggtttggca
     8101 agctagctgc agtaacgcca ttttgcaagg catggaaaaa taccaaacca agaatagaga
     8161 agttcagatc aagggcgggt acatgaaaat agctaacgtt gggccaaaca ggatatctgc
     8221 ggtgagcagt ttcggccccg gcccggggcc aagaacagat ggtcaccgca gtttcggccc
     8281 cggcccgagg ccaagaacag atggtcccca gatatggccc aaccctcagc agtttcttaa
     8341 gacccatcag atgtttccag gctcccccaa ggacctgaaa tgaccctgcg ccttatttga
     8401 attaaccaat cagcctgctt ctcgcttctg ttcgcgcgct tctgcttccc gagctctata
     8461 aaagagctca caacccctca ctcggcgcgc cagtcctccg acagactgag tcgcccgggg
     8521 gggatctgga gctctcgaga attctcacgc gtgccaccat gcctgctgct aaacgtgtta
     8581 agcttgatgg gggtagcccg gcagccaaga gagtcaaact cgacggcggc tccccagcgg
     8641 caaaaagggt gaaactagac gggggtagcc ccgccgcgaa gcgtgtaaag ctggatggag
     8701 gatcgcctgc ggctaagcga gtcaaattag acggtgggtc accggctgca aagcgcgtca
     8761 aattggatgc tagcggcaca cagttcgagg gctttaccaa cctgtatcag gtgagcaaga
     8821 cactgcggtt tgagctgatc ccacagggca agaccctgaa gcacatccag gagcagggct
     8881 tcatcgagga ggacaaggcc cgcaatgatc actacaagga gctgaagccc atcatcgatc
     8941 ggatctacaa gacctatgcc gaccagtgcc tgcagctggt gcagctggat tgggagaacc
     9001 tgagcgccgc catcgactcc tatagaaagg agaaaaccga ggagacaagg aacgccctga
     9061 tcgaggagca ggccacatat cgcaatgcca tccacgacta cttcatcggc cggacagaca
     9121 acctgaccga tgccatcaat aagagacacg ccgagatcta caagggcctg ttcaaggccg
     9181 agctgtttaa tggcaaggtg ctgaagcagc tgggcaccgt gaccacaacc gagcacgaga
     9241 acgccctgct gcggagcttc gacaagttta caacctactt ctccggcttt tatagaaaca
     9301 ggaagaacgt gttcagcgcc gaggatatca gcacagccat cccacaccgc atcgtgcagg
     9361 acaacttccc caagtttaag gagaattgtc acatcttcac acgcctgatc accgccgtgc
     9421 ccagcctgcg ggagcacttt gagaacgtga agaaggccat cggcatcttc gtgagcacct
     9481 ccatcgagga ggtgttttcc ttcccttttt ataaccagct gctgacacag acccagatcg
     9541 acctgtataa ccagctgctg ggaggaatct ctcgggaggc aggcaccgag aagatcaagg
     9601 gcctgaacga ggtgctgaat ctggccatcc agaagaatga tgagacagcc cacatcatcg
     9661 cctccctgcc acacagattc atccccctgt ttaagcagat cctgtccgat aggaacaccc
     9721 tgtctttcat cctggaggag tttaagagcg acgaggaagt gatccagtcc ttctgcaagt
     9781 acaagacact gctgagaaac gagaacgtgc tggagacagc cgaggccctg tttaacgagc
     9841 tgaacagcat cgacctgaca cacatcttca tcagccacaa gaagctggag acaatcagca
     9901 gcgccctgtg cgaccactgg gatacactga ggaatgccct gtatgagcgg agaatctccg
     9961 agctgacagg caagatcacc aagtctgcca aggagaaggt gcagcgcagc ctgaagcacg
    10021 aggatatcaa cctgcaggag atcatctctg ccgcaggcaa ggagctgagc gaggccttca
    10081 agcagaaaac cagcgagatc ctgtcccacg cacacgccgc cctggatcag ccactgccta
    10141 caaccctgaa gaagcaggag gagaaggaga tcctgaagtc tcagctggac agcctgctgg
    10201 gcctgtacca cctgctggac tggtttgccg tggatgagtc caacgaggtg gaccccgagt
    10261 tctctgcccg gctgaccggc atcaagctgg agatggagcc ttctctgagc ttctacaaca
    10321 aggccagaaa ttatgccacc aagaagccct actccgtgga gaagttcaag ctgaactttc
    10381 agatgcctac actggccaga ggctgggacg tgaatagaga gaagaacaat ggcgccatcc
    10441 tgtttgtgaa gaacggcctg tactatctgg gcatcatgcc aaagcagaag ggcaggtata
    10501 aggccctgag cttcgagccc acagagaaaa ccagcgaggg ctttgataag atgtactatg
    10561 actacttccc tgatgccgcc aagatgatcc caaagtgcag cacccagctg aaggccgtga
    10621 cagcccactt tcagacccac acaaccccca tcctgctgtc caacaatttc atcgagcctc
    10681 tggagatcac aaaggagatc tacgacctga acaatcctga gaaggagcca aagaagtttc
    10741 agacagccta cgccaagaaa accggcgacc agaagggcta cagagaggcc ctgtgcaagt
    10801 ggatcgactt cacaagggat tttctgtcca agtataccaa gacaacctct atcgatctgt
    10861 ctagcctgcg gccatcctct cagtataagg acctgggcga gtactatgcc gagctgaatc
    10921 ccctgctgta ccacatcagc ttccagagaa tcgccgagaa ggagatcatg gatgccgtgg
    10981 agacaggcaa gctgtacctg ttccagatct ataacaagga ctttgccaag ggccaccacg
    11041 gcaagcctaa tctgcacaca ctgtattgga ccggcctgtt ttctccagag aacctggcca
    11101 agacaagcat caagctgaat ggccaggccg agctgttcta ccgccctaag tccaggatga
    11161 agaggatggc acaccggctg ggagagaaga tgctgaacaa gaagctgaag gatcagaaaa
    11221 ccccaatccc cgacaccctg taccaggagc tgtacgacta tgtgaatcac agactgtccc
    11281 acgacctgtc tgatgaggcc agggccctgc tgcccaacgt gatcaccaag gaggtgtctc
    11341 acgagatcat caaggatagg cgctttacca gcgacaagtt ctttttccac gtgcctatca
    11401 cactgaacta tcaggccgcc aattccccat ctaagttcaa ccagagggtg aatgcctacc
    11461 tgaaggagca ccccgagaca cctatcatcg gcatcgaccg gggcgagaga aacctgatct
    11521 atatcacagt gatcgactcc accggcaaga tcctggagca gcggagcctg aacaccatcc
    11581 agcagtttga ttaccagaag aagctggaca acagggagaa ggagagggtg gcagcaaggc
    11641 aggcctggtc tgtggtgggc acaatcaagg atctgaagca gggctatctg agccaggtca
    11701 tccacgagat cgtggacctg atgatccact accaggccgt ggtggtgctg gagaacctga
    11761 atttcggctt taagagcaag aggaccggca tcgccgagaa ggccgtgtac cagcagttcg
    11821 agaagatgct gatcgataag ctgaattgcc tggtgctgaa ggactatcca gcagagaaag
    11881 tgggaggcgt gctgaaccca taccagctga cagaccagtt cacctccttt gccaagatgg
    11941 gcacccagtc tggcttcctg ttttacgtgc ctgccccata tacatctaag atcgatcccc
    12001 tgaccggctt cgtggacccc ttcgtgtgga aaaccatcaa gaatcacgag agccgcaagc
    12061 acttcctgga gggcttcgac tttctgcact acgacgtgaa aaccggcgac ttcatcctgc
    12121 actttaagat gaacagaaat ctgtccttcc agaggggcct gcccggcttt atgcctgcat
    12181 gggatatcgt gttcgagaag aacgagacac agtttgacgc caagggcacc cctttcatcg
    12241 ccggcaagag aatcgtgcca gtgatcgaga atcacagatt caccggcaga taccgggacc
    12301 tgtatcctgc caacgagctg atcgccctgc tggaggagaa gggcatcgtg ttcagggatg
    12361 gctccaacat cctgccaaag ctgctggaga atgacgattc tcacgccatc gacacgatgg
    12421 tggccctgat ccgcagcgtg ctgcagatgg ccaactccaa tgccgccaca ggcgaggact
    12481 atatcaacag ccccgtgcgc gatctgaatg gcgtgtgctt cgactcccgg tttcagaacc
    12541 cagagtggcc aatggacgcc gatgccaatg gcgcctacca catcgccctg aagggccagc
    12601 tgctgctgaa tcacctgaag gagagcaagg atctgaagct gcagaacggc atctccaatc
    12661 aggactggct ggcctacatc caggagctgc gcaacagcag ggctgactat ccctatgacg
    12721 tgcccgatta tgccagcggc tcccccaaga agaagaggaa ggtgggatcc ggagggccga
    12781 gctctggcgc acccccacca agtggagggt ctcctgccgg gtccccaaca tctactgaag
    12841 aaggcaccag cgaatccgca acgcccgagt caggccctgg tacctccaca gaaccatctg
    12901 aaggtagtgc gcctggttcc ccagctggaa gccctacttc caccgaagaa ggcacgtcaa
    12961 ccgaaccaag tgaaggatct gcccctggga ccagcactga accatctgag atggatgcta
    13021 agtcactaac tgcctggtcc cggacactgg tgaccttcaa ggatgtattt gtggacttca
    13081 ccagggagga gtggaagctg ctggacactg ctcagcagat cgtgtacaga aatgtgatgc
    13141 tggagaacta taagaacctg gtttccttgg gttatcagct tactaagcca gatgtgatcc
    13201 tccggttgga gaagggagaa gagcccagcg gccgcgctac taacttcagc ctgctgaagc
    13261 aggctggaga cgtggaggag aaccctggac ctgtgtctaa gggcgaagag ctgattaagg
    13321 agaacatgca catgaagctg tacatggagg gcaccgtgga caaccatcac ttcaagtgca
    13381 catccgaggg cgaaggcaag ccctacgagg gcacccagac catgagaatc aaggtggtcg
    13441 agggcggccc tctccccttc gccttcgaca tcctggctac tagcttcctc tacggcagca
    13501 agaccttcat caaccacacc cagggcatcc ccgacttctt caagcagtcc ttccctgagg
    13561 gcttcacatg ggagagagtc accacatacg aagacggggg cgtgctgacc gctacccagg
    13621 acaccagcct ccaggacggc tgcctcatct acaacgtcaa gatcagaggg gtgaacttca
    13681 catccaacgg ccctgtgatg cagaagaaaa cactcggctg ggaggccttc accgagacgc
    13741 tgtaccccgc tgacggcggc ctggaaggca gaaacgacat ggccctgaag ctcgtgggcg
    13801 ggagccatct gatcgcaaac gccaagacca catatagatc caagaaaccc gctaagaacc
    13861 tcaagatgcc tggcgtctac tatgtggact acagactgga aagaatcaag gaggccaaca
    13921 acgagaccta cgtcgagcag cacgaggtgg cagtggccag atactgcgac ctccctagca
    13981 aactggggca caagcttaat taagcgatcg ccctgcaggc atgcaagctt gatatcaagc
    14041 ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
    14101 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
    14161 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
    14221 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
    14281 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
    14341 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
    14401 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
    14461 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
    14521 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
    14581 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
    14641 gataccgtcg acctcgaggg aattaattcg agctcggtac ctttaagacc aatgacttac
    14701 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
    14761 cactcccaac gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag
    14821 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
    14881 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
    14941 tccctcagac ccttttagtc agtgtggaaa atctctagca gc
//
