LOCUS       sequence-426480-       11428 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  piggyBac expression of multiAsCas12a.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11428)
  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 11428)
  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: pNS35 (multiAsCas12a piggyBac)
FEATURES             Location/Qualifiers
     source          1..11428
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        147..526
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          871..1887
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1811..1833
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1895..1914
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1956..1982
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             1992..2018
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2028..2054
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2064..2090
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2100..2126
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2136..2162
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             2172..6089
                     /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             6102..6128
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             6138..6158
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6177..6233
                     /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             6234..6941
                     /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    6981..7569
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7034..7054)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7452..7463)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(7608..7625)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7614..7838
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   8192..8226
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(8313..8332)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8314..8332)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8339..8356)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8339..8355)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8348..8370)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(8497..8952)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8634..8655)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8846..8865
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        8979..9083
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9084..9152,9153..9944)
                     /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(9302..9321)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10115..10703
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10604..10623
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     10857..10874
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    10991..11012
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11027..11032,11033..11050,11051..11057)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11065..11081
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     11070..11092
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11089..11105
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11089..11105
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11124..11144
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11126..11144
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   11208..11270
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 aagtaacaaa acttttaaac attctctctt ttacaaaaat aaacttattt tgtactttaa
       61 aaacagtcat gttgtattat aaaataagta attagcttaa cctatacata atagaaacaa
      121 attatactta ttaaccggtc atggtcgaca ttgattattg actagttatt aatagtaatc
      181 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      241 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      301 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      361 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      421 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      481 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc
      541 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt
      601 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggg ggccgccggg
      661 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg ggggcgcgcg
      721 ccaggcgggg cggggcgggg cgaggggcgg ggcggggcga ggcggagagg tgcggcggca
      781 gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg
      841 ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgcgctgcct tcgccccgtg
      901 ccccgctccg ccgccgcctc gcgccgcccg ccccggctct gactgaccgc gttactccca
      961 caggtgagcg ggcgggacgg cccttctcct ccgggctgta attagcgctt ggtttaatga
     1021 cggcttgttt cttttctgtg gctgcgtgaa agccttgagg ggctccggga gggccctttg
     1081 tgcgggggga gcggctcggg gggtgcgtgc gtgtgtgtgt gcgtggggag cgccgcgtgc
     1141 ggctccgcgc tgcccggcgg ctgtgagcgc tgcgggcgcg gcgcggggct ttgtgcgctc
     1201 cgcagtgtgc gcgaggggag cgcggccggg ggcggtgccc cgcggtgcgg ggggggctgc
     1261 gaggggaaca aaggctgcgt gcggggtgtg tgcgtggggg ggtgagcagg gggtgtgggc
     1321 gcgtcggtcg ggctgcaacc ccccctgcac ccccctcccc gagttgctga gcacggcccg
     1381 gcttcgggtg cggggctccg tacggggcgt ggcgcggggc tcgccgtgcc gggcgggggg
     1441 tggcggcagg tgggggtgcc gggcggggcg gggccgcctc gggccgggga gggctcgggg
     1501 gaggggcgcg gcggcccccg gagcgccggc ggctgtcgag gcgcggcgag ccgcagccat
     1561 tgccttttat ggtaatcgtg cgagagggcg cagggacttc ctttgtccca aatctgtgcg
     1621 gagccgaaat ctgggaggcg ccgccgcacc ccctctagcg ggcgcggggc gaagcggtgc
     1681 ggcgccggca ggaaggaaat gggcggggag ggccttcgtg cgtcgccgcg ccgccgtccc
     1741 cttctccctc tccagcctcg gggctgtccg cggggggacg gctgccttcg ggggggacgg
     1801 ggcagggcgg ggttcggctt ctggcgtgtg accggcggct ctagagcctc tgctaaccat
     1861 gttcatgcct tcttcttttt cctacagctc ctgggcaacg tgctggttat tgtgctgtct
     1921 catcattttg gcaaagaatt gctagcgcca ccatgcctgc tgctaaacgt gttaagcttg
     1981 atgggggtag cccggcagcc aagagagtca aactcgacgg cggctcccca gcggcaaaaa
     2041 gggtgaaact agacgggggt agccccgccg cgaagcgtgt aaagctggat ggaggatcgc
     2101 ctgcggctaa gcgagtcaaa ttagacggtg ggtcaccggc tgcaaagcgc gtcaaattgg
     2161 atgctagcgg cacacagttc gagggcttta ccaacctgta tcaggtgagc aagacactgc
     2221 ggtttgagct gatcccacag ggcaagaccc tgaagcacat ccaggagcag ggcttcatcg
     2281 aggaggacaa ggcccgcaat gatcactaca aggagctgaa gcccatcatc gatcggatct
     2341 acaagaccta tgccgaccag tgcctgcagc tggtgcagct ggattgggag aacctgagcg
     2401 ccgccatcga ctcctataga aaggagaaaa ccgaggagac aaggaacgcc ctgatcgagg
     2461 agcaggccac atatcgcaat gccatccacg actacttcat cggccggaca gacaacctga
     2521 ccgatgccat caataagaga cacgccgaga tctacaaggg cctgttcaag gccgagctgt
     2581 ttaatggcaa ggtgctgaag cagctgggca ccgtgaccac aaccgagcac gagaacgccc
     2641 tgctgcggag cttcgacaag tttacaacct acttctccgg cttttataga aacaggaaga
     2701 acgtgttcag cgccgaggat atcagcacag ccatcccaca ccgcatcgtg caggacaact
     2761 tccccaagtt taaggagaat tgtcacatct tcacacgcct gatcaccgcc gtgcccagcc
     2821 tgcgggagca ctttgagaac gtgaagaagg ccatcggcat cttcgtgagc acctccatcg
     2881 aggaggtgtt ttccttccct ttttataacc agctgctgac acagacccag atcgacctgt
     2941 ataaccagct gctgggagga atctctcggg aggcaggcac cgagaagatc aagggcctga
     3001 acgaggtgct gaatctggcc atccagaaga atgatgagac agcccacatc atcgcctccc
     3061 tgccacacag attcatcccc ctgtttaagc agatcctgtc cgataggaac accctgtctt
     3121 tcatcctgga ggagtttaag agcgacgagg aagtgatcca gtccttctgc aagtacaaga
     3181 cactgctgag aaacgagaac gtgctggaga cagccgaggc cctgtttaac gagctgaaca
     3241 gcatcgacct gacacacatc ttcatcagcc acaagaagct ggagacaatc agcagcgccc
     3301 tgtgcgacca ctgggataca ctgaggaatg ccctgtatga gcggagaatc tccgagctga
     3361 caggcaagat caccaagtct gccaaggaga aggtgcagcg cagcctgaag cacgaggata
     3421 tcaacctgca ggagatcatc tctgccgcag gcaaggagct gagcgaggcc ttcaagcaga
     3481 aaaccagcga gatcctgtcc cacgcacacg ccgccctgga tcagccactg cctacaaccc
     3541 tgaagaagca ggaggagaag gagatcctga agtctcagct ggacagcctg ctgggcctgt
     3601 accacctgct ggactggttt gccgtggatg agtccaacga ggtggacccc gagttctctg
     3661 cccggctgac cggcatcaag ctggagatgg agccttctct gagcttctac aacaaggcca
     3721 gaaattatgc caccaagaag ccctactccg tggagaagtt caagctgaac tttcagatgc
     3781 ctacactggc cagaggctgg gacgtgaata gagagaagaa caatggcgcc atcctgtttg
     3841 tgaagaacgg cctgtactat ctgggcatca tgccaaagca gaagggcagg tataaggccc
     3901 tgagcttcga gcccacagag aaaaccagcg agggctttga taagatgtac tatgactact
     3961 tccctgatgc cgccaagatg atcccaaagt gcagcaccca gctgaaggcc gtgacagccc
     4021 actttcagac ccacacaacc cccatcctgc tgtccaacaa tttcatcgag cctctggaga
     4081 tcacaaagga gatctacgac ctgaacaatc ctgagaagga gccaaagaag tttcagacag
     4141 cctacgccaa gaaaaccggc gaccagaagg gctacagaga ggccctgtgc aagtggatcg
     4201 acttcacaag ggattttctg tccaagtata ccaagacaac ctctatcgat ctgtctagcc
     4261 tgcggccatc ctctcagtat aaggacctgg gcgagtacta tgccgagctg aatcccctgc
     4321 tgtaccacat cagcttccag agaatcgccg agaaggagat catggatgcc gtggagacag
     4381 gcaagctgta cctgttccag atctataaca aggactttgc caagggccac cacggcaagc
     4441 ctaatctgca cacactgtat tggaccggcc tgttttctcc agagaacctg gccaagacaa
     4501 gcatcaagct gaatggccag gccgagctgt tctaccgccc taagtccagg atgaagagga
     4561 tggcacaccg gctgggagag aagatgctga acaagaagct gaaggatcag aaaaccccaa
     4621 tccccgacac cctgtaccag gagctgtacg actatgtgaa tcacagactg tcccacgacc
     4681 tgtctgatga ggccagggcc ctgctgccca acgtgatcac caaggaggtg tctcacgaga
     4741 tcatcaagga taggcgcttt accagcgaca agttcttttt ccacgtgcct atcacactga
     4801 actatcaggc cgccaattcc ccatctaagt tcaaccagag ggtgaatgcc tacctgaagg
     4861 agcaccccga gacacctatc atcggcatcg accggggcga gagaaacctg atctatatca
     4921 cagtgatcga ctccaccggc aagatcctgg agcagcggag cctgaacacc atccagcagt
     4981 ttgattacca gaagaagctg gacaacaggg agaaggagag ggtggcagca aggcaggcct
     5041 ggtctgtggt gggcacaatc aaggatctga agcagggcta tctgagccag gtcatccacg
     5101 agatcgtgga cctgatgatc cactaccagg ccgtggtggt gctggagaac ctgaatttcg
     5161 gctttaagag caagaggacc ggcatcgccg agaaggccgt gtaccagcag ttcgagaaga
     5221 tgctgatcga taagctgaat tgcctggtgc tgaaggacta tccagcagag aaagtgggag
     5281 gcgtgctgaa cccataccag ctgacagacc agttcacctc ctttgccaag atgggcaccc
     5341 agtctggctt cctgttttac gtgcctgccc catatacatc taagatcgat cccctgaccg
     5401 gcttcgtgga ccccttcgtg tggaaaacca tcaagaatca cgagagccgc aagcacttcc
     5461 tggagggctt cgactttctg cactacgacg tgaaaaccgg cgacttcatc ctgcacttta
     5521 agatgaacag aaatctgtcc ttccagaggg gcctgcccgg ctttatgcct gcatgggata
     5581 tcgtgttcga gaagaacgag acacagtttg acgccaaggg cacccctttc atcgccggca
     5641 agagaatcgt gccagtgatc gagaatcaca gattcaccgg cagataccgg gacctgtatc
     5701 ctgccaacga gctgatcgcc ctgctggagg agaagggcat cgtgttcagg gatggctcca
     5761 acatcctgcc aaagctgctg gagaatgacg attctcacgc catcgacacg atggtggccc
     5821 tgatccgcag cgtgctgcag atggccaact ccaatgccgc cacaggcgag gactatatca
     5881 acagccccgt gcgcgatctg aatggcgtgt gcttcgactc ccggtttcag aacccagagt
     5941 ggccaatgga cgccgatgcc aatggcgcct accacatcgc cctgaagggc cagctgctgc
     6001 tgaatcacct gaaggagagc aaggatctga agctgcagaa cggcatctcc aatcaggact
     6061 ggctggccta catccaggag ctgcgcaaca gcagggctga ctatccctat gacgtgcccg
     6121 attatgccag cggctccccc aagaagaaga ggaaggtggg atccggaggg ggccgcgcta
     6181 ctaacttcag cctgctgaag caggctggag acgtggagga gaaccctgga cctgtgtcta
     6241 agggcgaaga gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtgg
     6301 acaaccatca cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga
     6361 ccatgagaat caaggtggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta
     6421 ctagcttcct ctacggcagc aagaccttca tcaaccacac ccagggcatc cccgacttct
     6481 tcaagcagtc cttccctgag ggcttcacat gggagagagt caccacatac gaagacgggg
     6541 gcgtgctgac cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca
     6601 agatcagagg ggtgaacttc acatccaacg gccctgtgat gcagaagaaa acactcggct
     6661 gggaggcctt caccgagacg ctgtaccccg ctgacggcgg cctggaaggc agaaacgaca
     6721 tggccctgaa gctcgtgggc gggagccatc tgatcgcaaa cgccaagacc acatatagat
     6781 ccaagaaacc cgctaagaac ctcaagatgc ctggcgtcta ctatgtggac tacagactgg
     6841 aaagaatcaa ggaggccaac aacgagacct acgtcgagca gcacgaggtg gcagtggcca
     6901 gatactgcga cctccctagc aaactggggc acaagcttaa ttaattaatt aatgatcata
     6961 cagtgctctg agtacgctat aatcaacctc tggattacaa aatttgtgaa agattgactg
     7021 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     7081 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     7141 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     7201 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     7261 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     7321 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     7381 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     7441 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     7501 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     7561 cctccccgca ggatccacta gagcggccgc ctagagctcg ctatcagcct cgactgtgcc
     7621 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     7681 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     7741 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     7801 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag
     7861 ctggggctcg actagagctt gcggaaccct taagcttttt tattatatat tagtcacgat
     7921 atctataaca agaaaatata tatataataa gttatcacgt aagtagaaca tgaaataaca
     7981 atataattat cgtatgagtt aaatcttaaa agtcacgtaa aagataatca tgcgtcattt
     8041 tgactcacgc ggtcgttata gttcaaaatc agtgacactt accgcattga caagcacgcc
     8101 tcacgggagc tccaagcggc gactgagatg tcctaaatgc acagcgacgg attcgcgcta
     8161 tttagaaaga gagagcaata tttcaagaat gcatgcgtca attttacgca gactatcttt
     8221 ctagggttaa aaaagatttg cgctttactc gacctaaact ttaaacacgt catagaatct
     8281 tcgtttgaca aaaaccacat tgtgttgggt cgccctatag tgagtcgtat tacaattcac
     8341 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     8401 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     8461 cttcccaaca gttgcgcagc ctgaatggcg aatggcaaat tgtaagcgtt aatattttgt
     8521 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     8581 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     8641 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     8701 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     8761 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     8821 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     8881 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     8941 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     9001 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     9061 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     9121 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     9181 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     9241 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     9301 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     9361 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     9421 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     9481 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     9541 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     9601 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     9661 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     9721 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     9781 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     9841 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     9901 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     9961 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
    10021 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
    10081 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
    10141 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
    10201 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
    10261 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
    10321 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
    10381 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
    10441 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
    10501 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
    10561 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
    10621 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
    10681 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
    10741 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
    10801 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
    10861 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt
    10921 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
    10981 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc
    11041 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
    11101 atgaccatga ttacgccaag cgcgcaatta accctcacta aagggaacct cccccggtat
    11161 tcacgacagc aggctgaata ataaaaaaat tagaaactat tatttaaccc tagaaagata
    11221 atcatattgt gacgtacgtt aaagataatc atgcgtaaaa ttgacgcatg tgttttatcg
    11281 gtctgtatat cgaggtttat ttattaattt gaatagatat taagttttat tatatttaca
    11341 cttacatact aataataaat tcaacaaaca atttatttat gtttatttat ttattaaaaa
    11401 aaaacaaaaa ctcaaaattt cttctata
//
