LOCUS       sequence-359523-       14756 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac Tet-ON plasmid for differentiating hiPSCs into skeletal 
            muscle cells via MYOD1 and shOCT4 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14756)
  TITLE     Characterization of iPSC lines for the iPSC Neurodegenerative 
            Disease Initiative (iNDI)
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 14756)
  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            "Unpublished"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: PB-TO-MYOD1-shOct4
FEATURES             Location/Qualifiers
     source          1..14756
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   21..83
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        1188..1565
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    1195..1213
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1231..1249
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1267..1285
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1303..1321
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1339..1357
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1375..1393
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1411..1429
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1474..1498
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1575..1584
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3099..3220
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3136..3155)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3190..3209
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3269..3285)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3293..3309
                     /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(3317..3323,3324..3341,3342..3347))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3362..3383
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     3809..3828
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    3827..3882
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     3928..3947
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(4038..4784)
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     primer_bind     complement(4816..4835)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          4843..5859
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     complement(4897..4919)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     enhancer        6139..6518
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     polyA_signal    6519..6567
                     /note="synthetic polyadenylation signal"
     misc_feature    6581..6672
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     6621..6640
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     protein_bind    complement(7884..7917)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        7918..8129
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          8130..9068
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     9025..9045
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             9091..9687
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(9091..9110)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     9587..9607
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             9697..9750
                     /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             9757..9783
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             9799..9825
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     regulatory      9847..9856
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             9853..10563
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRND
                     MALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     polyA_signal    10583..10807
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(10808..10841)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     repeat_region   11357..11391
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(11882..11898)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    11906..11922
                     /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(11930..11936,11937..11954,11955..11960))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11975..11996
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(12284..12872)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12364..12383)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13043..13834,13835..13903))
                     /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     13666..13685
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13904..14008)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(14034..14489)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(14171..14192)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     14383..14402
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     14616..14638
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     14630..14647
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     14631..14647
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 agaacacgca gctagattaa ccctagaaag ataatcatat tgtgacgtac gttaaagata
       61 atcatgtgta aaattgacgc atgtgtttta tcggtctgta tatcgaggtt tatttattaa
      121 tttgaataga tattaagttt tattatattt acacttacat actaataata aattcaacaa
      181 acaatttatt tatgtttatt tatttattaa aaaaaacaaa aactcaaaat ttcttctata
      241 aagtaacaaa acttttatga gggacagccc ccccccaaag cccccaggga tgtaattacg
      301 tccctccccc gctagggggc agcagcgagc cgcccggggc tccgctccgg tccggcgctc
      361 cccccgcatc cccgagccgg cagcgtgcgg ggacagcccg ggcacgggga aggtggcacg
      421 ggatcgcttt cctctgaacg cttctcgctg ctctttgagc ctgcagacac ctggggggat
      481 acggggaaaa ggcctccacg gccttaatta agggaggtgg tccctgcagt tacgccaatg
      541 ataacccccg ccagaaaaat cttagtagcc ttcccttttt gttttccgtg ccccaactcg
      601 gcggattgac tcggcccctt ccggaaacac ccgaatcaac ttctagtcaa attattgttc
      661 acgccgcaat gacccacccc tggcccgcgt ctgtggaact gacccctggt gtacaggaga
      721 gttcgctgct gaaagtggtc ccaaaggggt actagttttt aagctcccaa ctccccctcc
      781 cccagcgtct ggaggattcc acaccctcgc accgcagggg cgaggaagtg ggcggagtcc
      841 ggttttggcg ccagccgctg aggctgccaa gcagaaaagc caccgctgag gagactccgg
      901 tcactgtcct cgccccgcct cccccttccc tccccttggg gaccaccggg cgccacgccg
      961 cgaacggtaa gtgccgcggt cgtcggcgcc tccgccctcc ccctagggcc ccaattccca
     1021 gcgggcgcgg cgcgcggccc ctccccccgc cgggcgcgcg cccgctgccc cgcccttcgt
     1081 ggccgcccgg cgtgggcggt gccacccctc cccccggcgg ccccgcgcgc agctcccggc
     1141 tccctccccc ttcggatgtg gcttgagctg taggcgcgga gggtaccagt ttactcccta
     1201 tcagtgatag agaacgtatg aagagtttac tccctatcag tgatagagaa cgtatgcaga
     1261 ctttactccc tatcagtgat agagaacgta taaggagttt actccctatc agtgatagag
     1321 aacgtatgac cagtttactc cctatcagtg atagagaacg tatctacagt ttactcccta
     1381 tcagtgatag agaacgtata tccagtttac tccctatcag tgatagagaa cgtataagct
     1441 ttaggcgtgt acggtgggcg cctataaaag cagagctcgt ttagtgaacc gtcagatcgc
     1501 ctggagcaat tccacaacac ttttgtctta taccaacttt ccgtaccact tcctaccctc
     1561 gtaaaggatc cgccgccacc atggagctac tgtcgccacc gctccgcgac gtagacctga
     1621 cggcccccga cggctctctc tgctcctttg ccacaacgga cgacttctat gacgacccgt
     1681 gtttcgactc cccggacctg cgcttcttcg aagacctgga cccgcgcctg atgcacgtgg
     1741 gcgcgctcct gaaacccgaa gagcactcgc acttccccgc ggcggtgcac ccggccccgg
     1801 gcgcacgtga ggacgagcat gtgcgcgcgc ccagcgggca ccaccaggcg ggccgctgcc
     1861 tactgtgggc ctgcaaggcg tgcaagcgca agaccaccaa cgccgaccgc cgcaaggccg
     1921 ccaccatgcg cgagcggcgc cgcctgagca aagtaaatga ggcctttgag acactcaagc
     1981 gctgcacgtc gagcaatcca aaccagcggt tgcccaaggt ggagatcctg cgcaacgcca
     2041 tccgctatat cgagggcctg caggctctgc tgcgcgacca ggacgccgcg ccccctggcg
     2101 ccgcagccgc cttctatgcg ccgggcccgc tgcccccggg ccgcggcggc gagcactaca
     2161 gcggcgactc cgacgcgtcc agcccgcgct ccaactgctc cgacggcatg atggactaca
     2221 gcggcccccc gagcggcgcc cggcggcgga actgctacga aggcgcctac tacaacgagg
     2281 cgcccagcga acccaggccc gggaagagtg cggcggtgtc gagcctagac tgcctgtcca
     2341 gcatcgtgga gcgcatctcc accgagagcc ctgcggcgcc cgccctcctg ctggcggacg
     2401 tgccttctga gtcgcctccg cgcaggcaag aggctgccgc ccccagcgag ggagagagca
     2461 gcggcgaccc cacccagtca ccggacgccg ccccgcagtg ccctgcgggt gcgaacccca
     2521 acccgatata ccaggtgctc taagtttaaa ccaccggtgt tcgtttgaat gaggcttcag
     2581 tactttacag aatcgttgcc tgcacatctt ggaaacactt gctgggatta cttcttcagc
     2641 ttaactcaga ccccttaccc acccagcaac ccaaagtcta ctctctagtc cttagggagg
     2701 ttgtgggggc ggaaaggggg acggggctga atttcttcct tccccaaccc ccttcccttc
     2761 tcctccagat agatgcaaag ctgaatctcc cgccctgctc gctcagctga tctgtggctt
     2821 acagaaccac actcggacgg gattgagttt tgaactcgtc cgagagtggt tctgtgagtt
     2881 acatcagtcg gttttcgtcg agggccccaa tcgagctctc ccactcctac cctccccagt
     2941 gggactgccc cactgccccc tcccagatag ggcaaagtgg gtgcagacca aggaggacaa
     3001 gctgtgagtg gggttgcaga acaagtctgg agaaccctgc tttatgccgt ccgtctagca
     3061 cgagacatcg ataaactcgc gaatcgatcg cccaccccaa cttgtttatt gcagcttata
     3121 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     3181 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     3241 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     3301 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     3361 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     3421 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3481 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3541 gagcggtatc agctcactca aaggacggcc gcacatgtaa ggaaaatttt agggatgtta
     3601 aagaaaaaaa taacacaaaa caaaatataa aaaaaatcta acctcaagtc aaggcttttc
     3661 tatggaataa ggaatggaca gcagggggct gtttcatata ctgatgacct ctttatagcc
     3721 aacctttgtt catggcagcc agcatatggg catatgttgc caaactctaa accaaatact
     3781 cattctgatg ttttaaatga tttgccctcc catatgtcct tccgagtgag agacacaaaa
     3841 aattccaaca cactattgca atgaaaataa atttccttta ttagccagaa gtcagatgct
     3901 caaggggctt catgatgtcc ccataatttt tggcagaggg aaaaagatct cagtggtatt
     3961 tgtgagccag ggcattggcc acaccagtca ccaccttctg ataggcagcc tgcacctgag
     4021 gagtgaattc acgcgtttta gcctggcagc atatcgagat cgaagtcatc gagagcgtca
     4081 gcagggagca tatccagatc aaaatcatcc agggcatcgg cgggcagcat gtccaggtcg
     4141 aagtcgtcca gggcgtcggt gggtccgccg ctctcgcact tcagctgctt ctcgaggccg
     4201 cagatgatca gttccaggcc gaacaggaag gcgggctcgg cgccctgtct gtcgaacagc
     4261 tcgatggcct gcttcagcag ggggggcatg ctgtcggtgg tgggtgtctc tctttcctct
     4321 ttggcgacct ggtgttcctg ttcttccagc acgcagccca gggtgaagtg gcccacggcg
     4381 ctcagggcgt acagggcgtt ttccaggctg aagccctgct ggcacaggaa ggccagctgg
     4441 ttttccagtg tctcgtactg cttctcggtg ggtctggtgc ccaggtgcac tttggcgccg
     4501 tcccggtggc tcagcagggc gcatctgtag ctcttggcgt tgttccgcag gaagtcctgc
     4561 cagctctcgc cttccagagg gcagctgtgg gtgtggtgcc ggtccagcat ctcgatgggc
     4621 agggcgtcca gcagggcccg cttgttcttc acgtgccagt acagggtggg ctgttccacg
     4681 cccagtttct gggccagctt ccgggtggtc aggccctcga tgcccacgcc gttcagcagt
     4741 tccagggcgc tgttgatcac tttgctcttg tccagccggc tcatgtcgac attctttgcc
     4801 aaaatgatga gacagcacaa taaccagcac gttgcccagg agctgtagga aaaagaagaa
     4861 ggcatgaaca tggttagcag aggctctaga gccgccggtc acacgccaga agccgaaccc
     4921 cgccctgccc cgtccccccc gaaggcagcc gtccccccgc ggacagcccc gaggctggag
     4981 agggagaagg ggacggcggc gcggcgacgc acgaaggccc tccccgccca tttccttcct
     5041 gccggcgccg caccgcttcg ccccgcgccc gctagagggg gtgcggcggc gcctcccaga
     5101 tttcggctcc gcacagattt gggacaaagg aagtccctgc gccctctcgc acgattacca
     5161 taaaaggcaa tggctgcggc tcgccgcgcc tcgacagccg ccggcgctcc gggggccgcc
     5221 gcgcccctcc cccgagccct ccccggcccg aggcggcccc gccccgcccg gcacccccac
     5281 ctgccgccac cccccgcccg gcacggcgag ccccgcgcca cgccccgtac ggagccccgc
     5341 acccgaagcc gggccgtgct cagcaactcg gggagggggg tgcagggggg gttgcagccc
     5401 gaccgacgcg cccacacccc ctgctcaccc ccccacgcac acaccccgca cgcagccttt
     5461 gttcccctcg cagccccccc cgcaccgcgg ggcaccgccc ccggccgcgc tcccctcgcg
     5521 cacactgcgg agcgcacaaa gccccgcgcc gcgcccgcag cgctcacagc cgccgggcag
     5581 cgcggagccg cacgcggcgc tccccacgca cacacacacg cacgcacccc ccgagccgct
     5641 ccccccgcac aaagggccct cccggagccc ctcaaggctt tcacgcagcc acagaaaaga
     5701 aacaagccgt cattaaacca agcgctaatt acagcccgga ggagaagggc cgtcccgccc
     5761 gctcacctgt gggagtaacg cggtcagtca gagccggggc gggcggcgcg aggcggcggc
     5821 ggagcggggc acggggcgaa ggcagcgcgc agcgactccc gcccgccgcg cgcttcgctt
     5881 tttatagggc cgccgccgcc gccgcctcgc cataaaagga aactttcgga gcgcgccgct
     5941 ctgattggct gccgccgcmc ctcyccsccy csccccsccc csccccycsc cccsccccsc
     6001 ccccccyggc gcgcgccccc cccccccccc cgcccccatc gctgcacaaa ataattaaaa
     6061 aataaataaa tacaaaattg ggggtgggga ggggggggag atggggagag tgaagcagaa
     6121 cgtggggctc acctcgacca tggtaatagc gatgactaat acgtagatgt actgccaagt
     6181 aggaaagtcc cataaggtca tgtactgggc ataatgccag gcgggccatt taccgtcatt
     6241 gacgtcaata gggggcgtac ttggcatatg atacacttga tgtactgcca agtgggcagt
     6301 ttaccgtaaa tactccaccc attgacgtca atggaaagtc cctattggcg ttactatggg
     6361 aacatacgtc attattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc
     6421 atttaccgta agttatgtaa cgcggaactc catatatggg ctatgaacta atgaccccgt
     6481 aattgattac tattaataac tagtcaataa tcaatgtcaa taaaatatct ttattttcat
     6541 tacatctgtg tgttggtttt ttgtgtgaat cgatagtact aacatacgct ctccatcaaa
     6601 acaaaacgaa acaaaacaaa ctagcaaaat aggctgtccc cagtgcaagt gcaggtgcca
     6661 gaacatttct ctgagctcac ggggacagcc cccccccaaa gcccccaggg atgtaattac
     6721 gtccctcccc cgctaggggg cagcagcgag ccgcccgggg ctccgctccg gtccggcgct
     6781 ccccccgcat ccccgagccg gcagcgtgcg gggacagccc gggcacgggg aaggtggcac
     6841 gggatcgctt tcctctgaac gcttctcgct gctctttgag cctgcagaca cctgggggga
     6901 tacggggaaa aagctttagg ctgaaagaga gatttagaat gacagaatca tagaacggcc
     6961 tgggttgcaa aggagcacag tgctcatcca gatccaaccc cctgctatgt gcagggtcat
     7021 caaccagcag cccaggctgc ccagagccac atccagcctg gccttgaatg cctgcaggga
     7081 tggggcatcc acagcctcct tgggcaacct gttcagtgcg tcaccaccct ctgggggaaa
     7141 aactgcctcc tcatatccaa cccaaacctc ccctgtctca gtgtaaagcc attccccctt
     7201 gtcctatcaa gggggagttt gctgtgacat tgttggtctg gggtgacaca tgtttgccaa
     7261 ttcagtgcat cacggagagg cagatcttgg ggataaggaa gtgcaggaca gcatggacgt
     7321 gggacatgca ggtgttgagg gctctgggac actctccaag tcacagcgtt cagaacagcc
     7381 ttaaggataa gaagatagga tagaaggaca aagagcaagt taaaacccag catggagagg
     7441 agcacaaaaa ggccacagac actgctggtc cctgtgtctg agcctgcatg tttgatggtg
     7501 tctggatgca agcagaaggg gtggaagagc ttgcctggag agatacagct gggtcagtag
     7561 gactgggaca ggcagctgga gaattgccat gtagatgttc atacaatcgt caaatcatga
     7621 aggctggaaa agccctccaa gatccccaag accaacccca acccacccac cgtgcccact
     7681 ggccatgtcc ctcagtgcca catccccaca gttcttcatc acctccaggg acggtgaccc
     7741 ccccacctcc gtgggcagct gtgccactgc agcaccgctc tttggagaag gtaaatcttg
     7801 ctaaatccag cccgaccctc ccctggcaca acgtaaggcc attatctctc atccaactcc
     7861 aggacggagt cagtgagaat attataactt cgtatagcat acattatacg aagttatggg
     7921 cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgaaccg
     7981 gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc
     8041 tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt
     8101 ttcgcaacgg gtttgccgcc agaacacagg taagtgccgt gtgtggttcc cgcgggcctg
     8161 gcctctttac gggttatggc ccttgcgtgc cttgaattac ttccacctgg ctgcagtacg
     8221 tgattcttga tcccgagctt cgggttggaa gtgggtggga gagttcgagg ccttgcgctt
     8281 aaggagcccc ttcgcctcgt gcttgagttg aggcctggcc tgggcgctgg ggccgccgcg
     8341 tgcgaatctg gtggcacctt cgcgcctgtc tcgctgcttt cgataagtct ctagccattt
     8401 aaaatttttg atgacctgct gcgacgcttt ttttctggca agatagtctt gtaaatgcgg
     8461 gccaagatct gcacactggt atttcggttt ttggggccgc gggcggcgac ggggcccgtg
     8521 cgtcccagcg cacatgttcg gcgaggcggg gcctgcgagc gcggccaccg agaatcggac
     8581 gggggtagtc tcaagctggc cggcctgctc tggtgcctgg cctcgcgccg ccgtgtatcg
     8641 ccccgccctg ggcggcaagg ctggcccggt cggcaccagt tgcgtgagcg gaaagatggc
     8701 cgcttcccgg ccctgctgca gggagctcaa aatggaggac gcggcgctcg ggagagcggg
     8761 cgggtgagtc acccacacaa aggaaaaggg cctttccgtc ctcagccgtc gcttcatgtg
     8821 actccacgga gtaccgggcg ccgtccaggc acctcgatta gttctcgcgc ttttggagta
     8881 cgtcgtcttt aggttggggg gaggggtttt atgcgatgga gtttccccac actgagtggg
     8941 tggagactga agttaggcca gcttggcact tgatgtaatt ctccttggaa tttgcccttt
     9001 ttgagtttgg atcttggttc attctcaagc ctcagacagt ggttcaaagt ttttttcttc
     9061 catttcaggt gtcgtgattt aaatcccacc atgaccgagt acaagcccac ggtgcgcctc
     9121 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     9181 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     9241 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     9301 gccgccgtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     9361 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     9421 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggagtc
     9481 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     9541 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     9601 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     9661 tggtgcatga cccgcaagcc cggtgccgga tcgggagagg gcagaggaag tctgctaaca
     9721 tgcggtgacg tcgaggagaa tcctggccca ccggtccctg ccgctaaacg ggtcaaactc
     9781 gacggcggtt ctggaggtcc tgctgcaaag cgcgttaagt tggatggagg gtcaggggga
     9841 gccaccgcca ccatggtgtc taagggcgaa gagctgatta aggagaacat gcacatgaag
     9901 ctgtacatgg agggcaccgt ggacaaccat cacttcaagt gcacatccga gggcgaaggc
     9961 aagccctacg agggcaccca gaccatgaga atcaaggtgg tcgagggcgg ccctctcccc
    10021 ttcgccttcg acatcctggc tactagcttc ctctacggca gcaagacctt catcaaccac
    10081 acccagggca tccccgactt cttcaagcag tccttccctg agggcttcac atgggagaga
    10141 gtcaccacat acgaagacgg gggcgtgctg accgctaccc aggacaccag cctccaggac
    10201 ggctgcctca tctacaacgt caagatcaga ggggtgaact tcacatccaa cggccctgtg
    10261 atgcagaaga aaacactcgg ctgggaggcc ttcaccgaga cgctgtaccc cgctgacggc
    10321 ggcctggaag gcagaaacga catggccctg aagctcgtgg gcgggagcca tctgatcgca
    10381 aacgccaaga ccacatatag atccaagaaa cccgctaaga acctcaagat gcctggcgtc
    10441 tactatgtgg actacagact ggaaagaatc aaggaggcca acaacgagac ctacgtcgag
    10501 cagcacgagg tggcagtggc cagatactgc gacctcccta gcaaactggg gcacaagctt
    10561 aattaggcgg ccgctaagaa ttctgtgcct tctagttgcc agccatctgt tgtttgcccc
    10621 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
    10681 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
    10741 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
    10801 tctatggata acttcgtata gcatacatta tacgaagtta tttggtatgg ctttttcccc
    10861 gtatcccccc aggtgtctgc aggctcaaag agcagcgaga agcgttcaga ggaaagcgat
    10921 cccgtgccac cttccccgtg cccgggctgt ccccgcacgc tgccggctcg gggatgcggg
    10981 gggagcgccg gaccggagcg gagccccggg cggctcgctg ctgcccccta gcgggggagg
    11041 gacgtaatta catccctggg ggctttgggg gggggctgtc cctgattcta taacaagaaa
    11101 atatatatat aataagttat cacgtaagta gaacatgaaa taacaatata attatcgtat
    11161 gagttaaatc ttaaaagtca cgtaaaagat aatcatgcgt cattttgact cacgcggtcg
    11221 ttatagttca aaatcagtga cacttaccgc attgacaagc acgcctcacg ggagctccaa
    11281 gcggcgactg agatgtccta aatgcacagc gacggattcg cgctatttag aaagagagag
    11341 caatatttca agaatgcatg cgtcaatttt acgcagacta tctttctagg gttaatctag
    11401 ctgcatcagg atcatatcgt cgggtctttt ttccggctca gtcatcgccc aagctggcgc
    11461 tatctgggca tcggggagga agaagcccgt gccttttccc gcgaggttga agcggcatgg
    11521 aaagagtttg ccgaggatga ctgctgctgc attgacgttg agcgaaaacg cacgtttacc
    11581 atgatgattc gggaaggtgt ggccatgcac gcctttaacg gtgaactgtt cgttcaggcc
    11641 acctgggata ccagttcgtc gcggcttttc cggacacagt tccggatggt cagcccgaag
    11701 cgcatcagca acccgaacaa taccggcgac agccggaact gccgtgccgg tgtgcagatt
    11761 aatgacagcg gtgcggcgct gggatattac gtcagcgagg acgggtatcc tggctggatg
    11821 ccgcagaaat ggacatggat accccgtgag ttacccggcg ggcgcgcttg gcgtaatcat
    11881 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
    11941 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
    12001 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
    12061 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
    12121 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
    12181 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
    12241 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
    12301 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
    12361 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
    12421 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
    12481 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
    12541 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
    12601 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
    12661 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
    12721 gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
    12781 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
    12841 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
    12901 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
    12961 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
    13021 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
    13081 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
    13141 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
    13201 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
    13261 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
    13321 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
    13381 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
    13441 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
    13501 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
    13561 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
    13621 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
    13681 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
    13741 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
    13801 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
    13861 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
    13921 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
    13981 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt
    14041 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
    14101 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
    14161 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
    14221 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
    14281 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
    14341 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
    14401 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
    14461 cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg
    14521 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
    14581 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
    14641 ggccagtgag cgcgcctcgt tcattcacgt ttttgaaccc gtggaggacg ggcagactcg
    14701 cggtgcaaat gtgttttaca gcgtgatgga gcagatgaag atgctcgaca cgctgc
//
