LOCUS       sequence-486073-       13767 bp    DNA     circular SYN 16-SEP-2025
DEFINITION  Mammalian expression of dHyperLbCas12a-KRAB with Blasticidin 
            resistance selectable marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13767)
  AUTHORS   Melore SM, Hamilton MC, Reddy TE
  TITLE     HyperCas12a enables highly-multiplexed epigenome editing screens.
  JOURNAL   bioRxiv [Preprint]. 2024 Jul 9:2024.07.08.602263. doi: 
            10.1101/2024.07.08.602263.
  PUBMED    39026853
REFERENCE   2  (bases 1 to 13767)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 16, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Jul 9:2024.07.08.602263. doi: 
            10.1101/2024.07.08.602263."
COMMENT     Sequence Label: SM266_EF1A-dHyperLbCas12a-KRAB-T2A-BlastR
FEATURES             Location/Qualifiers
     source          1..13767
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2630..3808
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2861..3799
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3756..3776
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3824..7507
                     /codon_start=1
                     /product="CRISPR-associated protein from Lachnospiraceae 
                     bacterium ND2006 (Zetsche et al., 2015)"
                     /label=LbCpf1
                     /translation="MSKLEKFTNCYSLSKTLRFKAIPVGKTQENIDNKRLLVEDEKRAE
                     DYKGVKKLLDRYYLSFINDVLHSIKLKNLNNYISLFRKKTRTEKENKELENLEINLRKE
                     IAKAFKGNEGYKSLFKKDIIETILPEFLDDKDEIALVNSFNGFTTAFTGFFRNRENMFS
                     EEAKSTSIAFRCINENLTRYISNMDIFEKVDAIFDKHEVQEIKEKILNSDYDVEDFFEG
                     EFFNFVLTQEGIRVYNAIIGGFVTESGEKIKGLNEYINLYNQKTKQKLPKFKPLYKQVL
                     SDRESLSFYGRGYTSDEEVLEVFRNTLNKNSEIFSSIKKLEKLFKNFDEYSSAGIFVKN
                     GPAISTISKRIFGEWNVIRDKWNAEYDDIHLKKKAVVTEKYEDDRRKSFKKIGSFSLEQ
                     LQEYADADLSVVEKLKEIIIQKVDEIYKVYGSSEKLFDADFVLEKSLKKNDAVVAIMKD
                     LLDSVKSFENYIKAFFGEGKETNRDESFYGDFVLAYDILLKVDHIYDAIRNYVTQKPYS
                     KDKFKLYFQNPQFMGGWDKDKETDYRATILRYGSKYYLAIMDKKYAKCLQKIDKDDVNG
                     NYEKINYKLLPGPNKMLPKVFFSKKWMAYYNPSEDIQKIYKNGTFKKGDMFNLNDCHKL
                     IDFFKDSISRYPKWSNAYDFNFSETEKYKDIAGFYREVEEQGYKVSFESASKKEVDKLV
                     EEGKLYMFQIYNKDFSDKSHGTPNLHTMYFKLLFDENNHGQIRLSGGAELFMRRASLKK
                     EELVVHPANSPIANKNPDNPKKTTTLSYDVYKDKRFSEDQYELHIPIAINKCPKNIFKI
                     NTEVRVLLKHDDNPYVIGIARGERNLLYIVVVDGKGNIVEQYSLNEIINNFNGIRIKTD
                     YHSLLDKKEKERFEARQNWTSIENIKELKAGYISQVVHKICELVEKYDAVIALEDLNSG
                     FKNSRVKVEKQVYQKFEKMLIDKLNYMVDKKSNPCATGGALKGYQITNKFESFKSMSTQ
                     NGFIFYIPAWLTSKIDPSTGFVNLLKTKYTSIADSKKFISSFDRIMYVPEEDLFEFALD
                     YKNFSRTDADYIKKWKLYSYGNRIRIFRNPKKNNVFDWEEVCLTSAYKELFNKYGINYQ
                     QGDIRALLCEQSDKAFYSSFMALMSLMLQMRNSITGRTDVDFLISPVKNSDGIFYDSRN
                     YEAQENAILPKNADANGAYNIARKVLWAIGQFKKAEDEKLDKVKIAISNKEWLEYAQTS
                     VKH"
     CDS             7514..7540
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             7520..7546
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             7559..7585
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             7592..7618
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             7682..7876
                     /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="RTLVTFKDVFVDFTREEWKLLDTAQQILYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEPWLV"
     CDS             7943..7963
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7970..8023
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             8030..8425
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    8459..9047
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8512..8532)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8930..8941)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9050..9066)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9051..9067)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9572..9752
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9778..9795)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9784..10008
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10054..10482
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10141..10160)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10351..10372
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10491..10511)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10496..10825
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10676..10811
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10738..10757
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10873..10920
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10939..11313
                     /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    11443..11564
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11480..11499)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11534..11553
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11613..11629)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11613..11629)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11626..11648)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11637..11653
                     /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(11661..11667,11668..11685,11686..11691))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11706..11727
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11844..11861)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12015..12603)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12095..12114)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12774..13565,13566..13634))
                     /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     13397..13416
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13635..13739)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta acccgtgtcg gctccagatc tggcctccgg gctccggtgc
     2641 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
     2701 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
     2761 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
     2821 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
     2881 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg
     2941 gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag
     3001 gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg
     3061 gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc
     3121 tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct
     3181 tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga
     3241 cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc
     3301 gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc
     3361 gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc
     3421 ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc
     3481 gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt
     3541 cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag
     3601 cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca
     3661 cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga
     3721 atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag
     3781 tttttttctt ccatttcagg tgtcgtgagc ttctagagcc accatgagca agctggagaa
     3841 gtttacaaac tgctactccc tgtctaagac cctgaggttc aaggccatcc ctgtgggcaa
     3901 gacccaggag aacatcgaca ataagcggct gctggtggag gacgagaaga gagccgagga
     3961 ttataagggc gtgaagaagc tgctggatcg ctactatctg tcttttatca acgacgtgct
     4021 gcacagcatc aagctgaaga atctgaacaa ttacatcagc ctgttccgga agaaaaccag
     4081 aaccgagaag gagaataagg agctggagaa cctggagatc aatctgcgga aggagatcgc
     4141 caaggccttc aagggcaacg agggctacaa gtccctgttt aagaaggata tcatcgagac
     4201 aatcctgcca gagttcctgg acgataagga cgagatcgcc ctggtgaaca gcttcaatgg
     4261 ctttaccaca gccttcaccg gcttctttcg caacagagag aatatgtttt ccgaggaggc
     4321 caagagcaca tccatcgcct tcaggtgtat caacgagaat ctgacccgct acatctctaa
     4381 tatggacatc ttcgagaagg tggacgccat ctttgataag cacgaggtgc aggagatcaa
     4441 ggagaagatc ctgaacagcg actatgatgt ggaggatttc tttgagggcg agttctttaa
     4501 ctttgtgctg acacaggagg gcatccgcgt gtataacgcc atcatcggcg gcttcgtgac
     4561 cgagagcggc gagaagatca agggcctgaa cgagtacatc aacctgtata atcagaaaac
     4621 caagcagaag ctgcctaagt ttaagccact gtataagcag gtgctgagcg atcgggagtc
     4681 tctgagcttc tacggccggg gctatacatc cgatgaggag gtgctggagg tgtttagaaa
     4741 caccctgaac aagaacagcg agatcttcag ctccatcaag aagctggaga agctgttcaa
     4801 gaattttgac gagtactcta gcgccggcat ctttgtgaag aacggccccg ccatcagcac
     4861 aatctccaag cgcatcttcg gcgagtggaa cgtgatccgg gacaagtgga atgccgagta
     4921 tgacgatatc cacctgaaga agaaggccgt ggtgaccgag aagtacgagg acgatcggag
     4981 aaagtccttc aagaagatcg gctccttttc tctggagcag ctgcaggagt acgccgacgc
     5041 cgatctgtct gtggtggaga agctgaagga gatcatcatc cagaaggtgg atgagatcta
     5101 caaggtgtat ggctcctctg agaagctgtt cgacgccgat tttgtgctgg agaagagcct
     5161 gaagaagaac gacgccgtgg tggccatcat gaaggacctg ctggattctg tgaagagctt
     5221 cgagaattac atcaaggcct tctttggcga gggcaaggag acaaacaggg acgagtcctt
     5281 ctatggcgat tttgtgctgg cctacgacat cctgctgaag gtggaccaca tctacgatgc
     5341 catccgcaat tatgtgaccc agaagcccta ctctaaggat aagttcaagc tgtattttca
     5401 gaaccctcag ttcatgggcg gctgggacaa ggataaggag acagactatc gggccaccat
     5461 cctgagatac ggctccaagt actatctggc catcatggat aagaagtacg ccaagtgcct
     5521 gcagaagatc gacaaggacg atgtgaacgg caattacgag aagatcaact ataagctgct
     5581 gcccggccct aataagatgc tgccaaaggt gttcttttct aagaagtgga tggcctacta
     5641 taaccccagc gaggacatcc agaagatcta caagaatggc acattcaaga agggcgatat
     5701 gtttaacctg aatgactgtc acaagctgat cgacttcttt aaggatagca tctcccggta
     5761 tccaaagtgg tccaatgcct acgatttcaa cttttctgag acagagaagt ataaggacat
     5821 cgccggcttt tacagagagg tggaggagca gggctataag gtgagcttcg agtctgccag
     5881 caagaaggag gtggataagc tggtggagga gggcaagctg tatatgttcc agatctataa
     5941 caaggacttt tccgataagt ctcacggcac acccaatctg cacaccatgt acttcaagct
     6001 gctgtttgac gagaacaatc acggacagat caggctgagc ggaggagcag agctgttcat
     6061 gaggcgcgcc tccctgaaga aggaggagct ggtggtgcac ccagccaact cccctatcgc
     6121 caacaagaat ccagataatc ccaagaaaac cacaaccctg tcctacgacg tgtataagga
     6181 taagaggttt tctgaggacc agtacgagct gcacatccca atcgccatca ataagtgccc
     6241 caagaacatc ttcaagatca atacagaggt gcgcgtgctg ctgaagcacg acgataaccc
     6301 ctatgtgatc ggcatcgcca ggggcgagcg caatctgctg tatatcgtgg tggtggacgg
     6361 caagggcaac atcgtggagc agtattccct gaacgagatc atcaacaact tcaacggcat
     6421 caggatcaag acagattacc actctctgct ggacaagaag gagaaggaga ggttcgaggc
     6481 ccgccagaac tggacctcca tcgagaatat caaggagctg aaggccggct atatctctca
     6541 ggtggtgcac aagatctgcg agctggtgga gaagtacgat gccgtgatcg ccctggagga
     6601 cctgaactct ggctttaaga atagccgcgt gaaggtggag aagcaggtgt atcagaagtt
     6661 cgagaagatg ctgatcgata agctgaacta catggtggac aagaagtcta atccttgtgc
     6721 aacaggcggc gccctgaagg gctatcagat caccaataag ttcgagagct ttaagtccat
     6781 gtctacccag aacggcttca tcttttacat ccctgcctgg ctgacatcca agatcgatcc
     6841 atctaccggc tttgtgaacc tgctgaaaac caagtatacc agcatcgccg attccaagaa
     6901 gttcatcagc tcctttgaca ggatcatgta cgtgcccgag gaggatctgt tcgagtttgc
     6961 cctggactat aagaacttct ctcgcacaga cgccgattac atcaagaagt ggaagctgta
     7021 ctcctacggc aaccggatca gaatcttccg gaatcctaag aagaacaacg tgttcgactg
     7081 ggaggaggtg tgcctgacca gcgcctataa ggagctgttc aacaagtacg gcatcaatta
     7141 tcagcagggc gatatcagag ccctgctgtg cgagcagtcc gacaaggcct tctactctag
     7201 ctttatggcc ctgatgagcc tgatgctgca gatgcggaac agcatcacag gccgcaccga
     7261 cgtggatttt ctgatcagcc ctgtgaagaa ctccgacggc atcttctacg atagccggaa
     7321 ctatgaggcc caggagaatg ccatcctgcc aaagaacgcc gacgccaatg gcgcctataa
     7381 catcgccaga aaggtgctgt gggccatcgg ccagttcaag aaggccgagg acgagaagct
     7441 ggataaggtg aagatcgcca tctctaacaa ggagtggctg gagtacgccc agaccagcgt
     7501 gaagcacgga tcctatccct atgacgtgcc cgattatgcc agcctgggca gcggctcccc
     7561 tgctgccaaa cgcgttaaac tagacgaaga tcctgccgcg aagcgcgtga aactggacga
     7621 cggcattggt agtgggagca acggcagcag cctcgatgct aagtcactga ctgcctggtc
     7681 ccggacactg gtgaccttca aggatgtgtt tgtggacttc accagggagg agtggaagct
     7741 gctggacact gctcagcaga tcctgtacag aaatgtgatg ctggagaact ataagaacct
     7801 ggtttccttg ggttatcagc ttactaagcc agatgtgatc ctccggttgg agaagggaga
     7861 agagccctgg ctggtggaga gagaaattca ccaagagacc catcctgatt cagagactgc
     7921 atttgaaatc aaatcatcag ttccgaaaaa gaaacgcaaa gttgctagcg agggcagagg
     7981 aagtcttcta acatgcggtg acgtggagga gaatcccggc cctggtacca tggccaagcc
     8041 tttgtctcaa gaagaatcca ccctcattga aagagcaacg gctacaatca acagcatccc
     8101 catctctgaa gactacagcg tcgccagcgc agctctctct agcgacggcc gcatcttcac
     8161 tggtgtcaat gtatatcatt ttactggggg accttgtgca gaactcgtgg tgctgggcac
     8221 tgctgctgct gcggcagctg gcaacctgac ttgtatcgtc gcgatcggaa atgagaacag
     8281 gggcatcttg agcccctgcg gacggtgccg acaggtgctt ctcgatctgc atcctgggat
     8341 caaagccata gtgaaggaca gtgatggaca gccgacggca gttgggattc gtgaattgct
     8401 gccctctggt tatgtgtggg agggcaccgg ttgagaattc gatatcaagc ttatcgataa
     8461 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     8521 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     8581 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     8641 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     8701 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     8761 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     8821 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     8881 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     8941 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     9001 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgtcg
     9061 acctcgagac ctagaaaaac atggagcaat cacaagtagc aatacagcag ctaccaatgc
     9121 tgattgtgcc tggctagaag cacaagagga ggaggaggtg ggttttccag tcacacctca
     9181 ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
     9241 aaagggggga ctggaagggc taattcactc ccaacgaaga caagatatcc ttgatctgtg
     9301 gatctaccac acacaaggct acttccctga ttggcagaac tacacaccag ggccagggat
     9361 cagatatcca ctgacctttg gatggtgcta caagctagta ccagttgagc aagagaaggt
     9421 agaagaagcc aatgaaggag agaacacccg cttgttacac cctgtgagcc tgcatgggat
     9481 ggatgacccg gagagagaag tattagagtg gaggtttgac agccgcctag catttcatca
     9541 catggcccga gagctgcatc cggactgtac tgggtctctc tggttagacc agatctgagc
     9601 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
     9661 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
     9721 acccttttag tcagtgtgga aaatctctag cagggcccgt ttaaacccgc tgatcagcct
     9781 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     9841 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     9901 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     9961 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
    10021 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
    10081 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
    10141 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
    10201 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
    10261 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
    10321 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
    10381 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
    10441 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
    10501 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
    10561 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
    10621 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
    10681 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
    10741 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
    10801 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
    10861 tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac
    10921 aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct caccgcgcgc
    10981 gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg ggacttcgtg
    11041 gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag cgcggtccag
    11101 gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct ggacgagctg
    11161 tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg gccggccatg
    11221 accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac
    11281 tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt cgattccacc
    11341 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
    11401 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
    11461 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
    11521 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
    11581 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
    11641 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
    11701 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
    11761 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
    11821 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
    11881 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
    11941 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
    12001 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
    12061 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
    12121 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
    12181 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
    12241 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
    12301 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
    12361 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
    12421 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
    12481 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
    12541 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
    12601 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
    12661 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
    12721 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
    12781 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
    12841 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
    12901 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
    12961 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
    13021 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
    13081 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
    13141 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
    13201 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
    13261 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
    13321 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
    13381 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
    13441 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
    13501 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
    13561 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
    13621 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
    13681 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
    13741 acatttcccc gaaaagtgcc acctgac
//
