LOCUS       sequence-230621-       14216 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral construct for inducible expression of dCas9-VP192 
            transcriptional activator and EGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14216)
  TITLE     Otonkoski plasmids 2018
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 14216)
  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: 
            "Unpublished"
COMMENT     Sequence Label: FU_tetO_dCas9-VP192-T2A-EGFP
FEATURES             Location/Qualifiers
     source          1..14216
                     /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"
     protein_bind    join(2610..2628,2629..2651,2652..2670,2671..2693,2694..2712,
                     2713..2735,2736..2754,2755..2777,2778..2796,2797..2819,
                     2820..2838,2839..2861,2862..2880)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        2913..2950
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2947..2971
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     3050..3069
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3050..3068
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3126..3152
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3156..3176
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(3186..3209,3210..3212,3213..5768,5769..5771,
                     5772..7286)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             join(3186..3209,3210..3212,3213..5699,5700..5702,
                     5703..7286)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7290..7310
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7347..7457
                     /codon_start=1
                     /product="3 tandem repeats of the minimal activation domain
                     of herpes simplex virus VP16 (Cheng et al., 2013)"
                     /label=VP48
                     /translation="DALDDFDLDMLPADALDDFDLDMLPADALDDFDLDML"
     CDS             7509..7619
                     /codon_start=1
                     /product="3 tandem repeats of the minimal activation domain
                     of herpes simplex virus VP16 (Cheng et al., 2013)"
                     /label=VP48
                     /translation="DALDDFDLDMLPADALDDFDLDMLPADALDDFDLDML"
     CDS             7671..7781
                     /codon_start=1
                     /product="3 tandem repeats of the minimal activation domain
                     of herpes simplex virus VP16 (Cheng et al., 2013)"
                     /label=VP48
                     /translation="DALDDFDLDMLPADALDDFDLDMLPADALDDFDLDML"
     CDS             7833..7943
                     /codon_start=1
                     /product="3 tandem repeats of the minimal activation domain
                     of herpes simplex virus VP16 (Cheng et al., 2013)"
                     /label=VP48
                     /translation="DALDDFDLDMLPADALDDFDLDMLPADALDDFDLDML"
     CDS             7983..8036
                     /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             join(8043..8045,8046..8048,8049..8759)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(8043..8045,8046..8048,8049..8759)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(8349..8368)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     8696..8717
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             8790..8831
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             8841..8858
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    8908..9496
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8961..8981)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9379..9390)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9499..9515)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9500..9516)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             10021..10201
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10227..10244)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10233..10457
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10503..10931
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10590..10609)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10800..10821
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10940..10960)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10945..11274
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      11125..11260
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11187..11206
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11322..11369
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11388..11762
                     /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    11892..12013
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11929..11948)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11983..12002
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(12062..12078)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12062..12078)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12075..12097)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12086..12102
                     /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(12110..12116,12117..12134,12135..12140))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12155..12176
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12293..12310)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12464..13052)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12544..12563)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13223..14014,14015..14083))
                     /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     13846..13865
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14084..14188)
                     /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 tggttatcga gtttaccact ccctatcagt gatagagaaa agtgaaagtc
     2641 gagtttacca ctccctatca gtgatagaga aaagtgaaag tcgagtttac cactccctat
     2701 cagtgataga gaaaagtgaa agtcgagttt accactccct atcagtgata gagaaaagtg
     2761 aaagtcgagt ttaccactcc ctatcagtga tagagaaaag tgaaagtcga gtttaccact
     2821 ccctatcagt gatagagaaa agtgaaagtc gagtttacca ctccctatca gtgatagaga
     2881 aaagtgaaag tcgagctcgg tacccgggtc gagtaggcgt gtacggtggg aggcctatat
     2941 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     3001 cctccataga agacaccggg accgatccag cctccgcggc cccgaattgt aatacgactc
     3061 actataggga gacccaagct ggctaggtaa gcttgatcaa caagtttgta cccggtgcca
     3121 ccatgtaccc atacgatgtt ccagattacg cttcgccgaa gaaaaagcgc aaggtcgaag
     3181 cgtccgacaa gaagtacagc atcggcctgg ccatcggcac caactctgtg ggctgggccg
     3241 tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc aacaccgacc
     3301 ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc gaaacagccg
     3361 aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag aaccggatct
     3421 gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc ttcttccaca
     3481 gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac cccatcttcg
     3541 gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatctac cacctgagaa
     3601 agaaactggt ggacagcacc gacaaggccg acctgcggct gatctatctg gccctggccc
     3661 acatgatcaa gttccggggc cacttcctga tcgagggcga cctgaacccc gacaacagcg
     3721 acgtggacaa gctgttcatc cagctggtgc agacctacaa ccagctgttc gaggaaaacc
     3781 ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc tgccagactg agcaagagca
     3841 gacggctgga aaatctgatc gcccagctgc ccggcgagaa gaagaatggc ctgttcggca
     3901 acctgattgc cctgagcctg ggcctgaccc ccaacttcaa gagcaacttc gacctggccg
     3961 aggatgccaa actgcagctg agcaaggaca cctacgacga cgacctggac aacctgctgg
     4021 cccagatcgg cgaccagtac gccgacctgt ttctggccgc caagaacctg tccgacgcca
     4081 tcctgctgag cgacatcctg agagtgaaca ccgagatcac caaggccccc ctgagcgcct
     4141 ctatgatcaa gagatacgac gagcaccacc aggacctgac cctgctgaaa gctctcgtgc
     4201 ggcagcagct gcctgagaag tacaaagaga ttttcttcga ccagagcaag aacggctacg
     4261 ccggctacat tgacggcgga gccagccagg aagagttcta caagttcatc aagcccatcc
     4321 tggaaaagat ggacggcacc gaggaactgc tcgtgaagct gaacagagag gacctgctgc
     4381 ggaagcagcg gaccttcgac aacggcagca tcccccacca gatccacctg ggagagctgc
     4441 acgccattct gcggcggcag gaagattttt acccattcct gaaggacaac cgggaaaaga
     4501 tcgagaagat cctgaccttc cgcatcccct actacgtggg ccctctggcc aggggaaaca
     4561 gcagattcgc ctggatgacc agaaagagcg aggaaaccat caccccctgg aacttcgagg
     4621 aagtggtgga caagggcgct tccgcccaga gcttcatcga gcggatgacc aacttcgata
     4681 agaacctgcc caacgagaag gtgctgccca agcacagcct gctgtacgag tacttcaccg
     4741 tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg aatgagaaag cccgccttcc
     4801 tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt caagaccaac cggaaagtga
     4861 ccgtgaagca gctgaaagag gactacttca agaaaatcga gtgcttcgac tccgtggaaa
     4921 tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac ataccacgat ctgctgaaaa
     4981 ttatcaagga caaggacttc ctggacaatg aggaaaacga ggacattctg gaagatatcg
     5041 tgctgaccct gacactgttt gaggacagag agatgatcga ggaacggctg aaaacctatg
     5101 cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg gcggagatac accggctggg
     5161 gcaggctgag ccggaagctg atcaacggca tccgggacaa gcagtccggc aagacaatcc
     5221 tggatttcct gaagtccgac ggcttcgcca acagaaactt catgcagctg atccacgacg
     5281 acagcctgac ctttaaagag gacatccaga aagcccaggt gtccggccag ggcgatagcc
     5341 tgcacgagca cattgccaat ctggccggca gccccgccat taagaagggc atcctgcaga
     5401 cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg gcacaagccc gagaacatcg
     5461 tgatcgaaat ggccagagag aaccagacca cccagaaggg acagaagaac agccgcgaga
     5521 gaatgaagcg gatcgaagag ggcatcaaag agctgggcag ccagatcctg aaagaacacc
     5581 ccgtggaaaa cacccagctg cagaacgaga agctgtacct gtactacctg cagaatgggc
     5641 gggatatgta cgtggaccag gaactggaca tcaaccggct gtccgactac gatgtcgacg
     5701 ccatcgtgcc tcagagcttt ctgaaggacg actccatcga caacaaggtg ctgaccagaa
     5761 gcgacaagaa ccggggcaag agcgacaacg tgccctccga agaggtcgtg aagaagatga
     5821 agaactactg gcggcagctg ctgaacgcca agctgattac ccagagaaag ttcgacaatc
     5881 tgaccaaggc cgagagaggc ggcctgagcg aactggataa ggccggcttc atcaagagac
     5941 agctggtgga aacccggcag atcacaaagc acgtggcaca gatcctggac tcccggatga
     6001 acactaagta cgacgagaat gacaagctga tccgggaagt gaaagtgatc accctgaagt
     6061 ccaagctggt gtccgatttc cggaaggatt tccagtttta caaagtgcgc gagatcaaca
     6121 actaccacca cgcccacgac gcctacctga acgccgtcgt gggaaccgcc ctgatcaaaa
     6181 agtaccctaa gctggaaagc gagttcgtgt acggcgacta caaggtgtac gacgtgcgga
     6241 agatgatcgc caagagcgag caggaaatcg gcaaggctac cgccaagtac ttcttctaca
     6301 gcaacatcat gaactttttc aagaccgaga ttaccctggc caacggcgag atccggaagc
     6361 ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt gtgggataag ggccgggatt
     6421 ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa tatcgtgaaa aagaccgagg
     6481 tgcagacagg cggcttcagc aaagagtcta tcctgcccaa gaggaacagc gataagctga
     6541 tcgccagaaa gaaggactgg gaccctaaga agtacggcgg cttcgacagc cccaccgtgg
     6601 cctattctgt gctggtggtg gccaaagtgg aaaagggcaa gtccaagaaa ctgaagagtg
     6661 tgaaagagct gctggggatc accatcatgg aaagaagcag cttcgagaag aatcccatcg
     6721 actttctgga agccaagggc tacaaagaag tgaaaaagga cctgatcatc aagctgccta
     6781 agtactccct gttcgagctg gaaaacggcc ggaagagaat gctggcctct gccggcgaac
     6841 tgcagaaggg aaacgaactg gccctgccct ccaaatatgt gaacttcctg tacctggcca
     6901 gccactatga gaagctgaag ggctcccccg aggataatga gcagaaacag ctgtttgtgg
     6961 aacagcacaa gcactacctg gacgagatca tcgagcagat cagcgagttc tccaagagag
     7021 tgatcctggc cgacgctaat ctggacaaag tgctgtccgc ctacaacaag caccgggata
     7081 agcccatcag agagcaggcc gagaatatca tccacctgtt taccctgacc aatctgggag
     7141 cccctgccgc cttcaagtac tttgacacca ccatcgaccg gaagaggtac accagcacca
     7201 aagaggtgct ggacgccacc ctgatccacc agagcatcac cggcctgtac gagacacgga
     7261 tcgacctgtc tcagctggga ggcgacagcc ccaagaagaa gagaaaggtg gaggccagcc
     7321 tcgagtgcga aagcggcggg ccggccgacg cccttgacga ttttgactta gacatgctcc
     7381 cagccgatgc ccttgacgac tttgaccttg atatgctgcc tgctgacgct cttgacgatt
     7441 ttgaccttga catgctcccc gggtcactac ccagcgctag cgtcgagtgc gaaagcggcg
     7501 ggccggccga cgcccttgac gattttgact tagacatgct cccagccgat gcccttgacg
     7561 actttgacct tgatatgctg cctgctgacg ctcttgacga ttttgacctt gacatgctcc
     7621 ccgggtcact acccagcgct agcgtcgagt gcgaaagcgg cgggccggcc gacgcccttg
     7681 acgattttga cttagacatg ctcccagccg atgcccttga cgactttgac cttgatatgc
     7741 tgcctgctga cgctcttgac gattttgacc ttgacatgct ccccgggtca ctacccagcg
     7801 ctagcgtcga gtgcgaaagc ggcgggccgg ccgacgccct tgacgatttt gacttagaca
     7861 tgctcccagc cgatgccctt gacgactttg accttgatat gctgcctgct gacgctcttg
     7921 acgattttga ccttgacatg ctccccgggt cactacccag cgctagcgtc gacggctccg
     7981 gagagggcag aggaagtctg ctaacatgcg gtgacgtcga ggagaatcct ggcccaatcg
     8041 atatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     8101 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     8161 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     8221 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     8281 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     8341 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     8401 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     8461 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     8521 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     8581 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     8641 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     8701 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaaag
     8761 tggttgatct agagggcccg cggttcgaag gtaagcctat ccctaaccct ctcctcggtc
     8821 tcgattctac gcgtaccggt catcatcacc atcaccattg agtttaaacc cgctgatcag
     8881 cctcaattcg atatcaagct tatcgataat caacctctgg attacaaaat ttgtgaaaga
     8941 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     9001 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     9061 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     9121 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     9181 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     9241 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     9301 aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     9361 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     9421 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     9481 tgggccgcct ccccgcatcg ataccgtcga cctcgagacc tagaaaaaca tggagcaatc
     9541 acaagtagca atacagcagc taccaatgct gattgtgcct ggctagaagc acaagaggag
     9601 gaggaggtgg gttttccagt cacacctcag gtacctttaa gaccaatgac ttacaaggca
     9661 gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc
     9721 caacgaagac aagatatcct tgatctgtgg atctaccaca cacaaggcta cttccctgat
     9781 tggcagaact acacaccagg gccagggatc agatatccac tgacctttgg atggtgctac
     9841 aagctagtac cagttgagca agagaaggta gaagaagcca atgaaggaga gaacacccgc
     9901 ttgttacacc ctgtgagcct gcatgggatg gatgacccgg agagagaagt attagagtgg
     9961 aggtttgaca gccgcctagc atttcatcac atggcccgag agctgcatcc ggactgtact
    10021 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
    10081 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
    10141 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
    10201 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
    10261 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
    10321 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
    10381 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
    10441 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
    10501 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
    10561 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
    10621 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
    10681 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
    10741 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
    10801 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
    10861 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
    10921 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
    10981 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
    11041 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
    11101 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
    11161 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
    11221 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
    11281 ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg
    11341 catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg gccaagttga
    11401 ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg
    11461 accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg gtccgggacg
    11521 acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac accctggcct
    11581 gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc gtgtccacga
    11641 acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg tgggggcggg
    11701 agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag gagcaggact
    11761 gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
    11821 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
    11881 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
    11941 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
    12001 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
    12061 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
    12121 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
    12181 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
    12241 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
    12301 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
    12361 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
    12421 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
    12481 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
    12541 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
    12601 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
    12661 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
    12721 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
    12781 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
    12841 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
    12901 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
    12961 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
    13021 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
    13081 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
    13141 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
    13201 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
    13261 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
    13321 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
    13381 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
    13441 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
    13501 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
    13561 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
    13621 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
    13681 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
    13741 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
    13801 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
    13861 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
    13921 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
    13981 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
    14041 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
    14101 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
    14161 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgac
//
