LOCUS       sequence-416546-       14598 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ngn2 mediated cortical neuron induction, SNCA(wt) expression, 
            Synapsin promoter BirA ligase expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14598)
  AUTHORS   Hallacli E, Kayatekin C, Nazeen S, Wang XH, Sheinkopf Z, Sathyakumar
            S, Sarkar S, Jiang X, Dong X, Di Maio R, Wang W, Keeney MT, Felsky 
            D, Sandoe J, Vahdatshoar A, Udeshi ND, Mani DR, Carr SA, Lindquist 
            S, De Jager PL, Bartel DP, Myers CL, Greenamyre JT, Feany MB, 
            Sunyaev SR, Chung CY, Khurana V
  TITLE     The Parkinson's disease protein alpha-synuclein is a modulator of 
            processing bodies and mRNA stability.
  JOURNAL   Cell. 2022 Jun 9;185(12):2035-2056.e33. doi: 
            10.1016/j.cell.2022.05.008.
  PUBMED    35688132
REFERENCE   2  (bases 1 to 14598)
  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"; doi: 
            "10.1016/j.cell.2022.05"; journalName: "Cell"; date: "2022-06-9- 9";
            volume: "185"; issue: "12"; pages: "2035-2056.e33"
COMMENT     Sequence Label: piggyBac_rtTA
            (4th_Gen)_SNCA_IRES_NGN2-2A-PURO...
FEATURES             Location/Qualifiers
     source          1..14598
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   132..166
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    719..943
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     932..949
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(972..1541)
                     /codon_start=1
                     /product="transcriptional activation domain from the human 
                     herpesvirus 4 (Epstein-Barr virus) replication and 
                     transcription activator Rta/BRLF1 (Hardwick et al., 1992; 
                     Chavez et al., 2015)"
                     /label=Rta AD
                     /translation="RDSREGMFLPKPEAGSAISDVFEGREVCQPKRIRPFHPPGSPWAN
                     RPLPASLAPTPTGPVHEPVGSLTPAPVPQPLDPAPAVTPEASHLLEDPDEETSQAVKAL
                     REMADTVIPQKEEAAICGQMDLSHPPPRGHLDELTTTLESMTEDLNLDSPLTPELNEIL
                     DTFLNDECLLHAMHISTGLSIFDTSLF"
     CDS             complement(1560..1916)
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALL"
     CDS             complement(2346..2366)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(2391..2540)
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             complement(3198..3251)
                     /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             complement(3261..3656)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     promoter        complement(3666..4176)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3729..3747)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     protein_bind    4888..4906
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4924..4942
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4998..5016
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5044..5062
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5091..5109
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5127..5145
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5266..5290
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      5294..5303
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(5724..5748)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    5800..6367
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    5817..6369
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(5961..5978)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     6188..6207
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             6381..6404
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7197..7250
                     /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             7257..7856
                     /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(7257..7276)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7753..7773
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7857..8445
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7910..7930)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8328..8339)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    8460..8936
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(8659..8678)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     promoter        8984..9431
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             join(9445..9795,9796..9798,9799..10407)
                     /codon_start=1
                     /product="promiscuous R118G mutant of the E. coli biotin 
                     protein ligase (Cronan, 2005)"
                     /label=BirA(R118G)
                     /note="also known as BirA* or BioID"
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             9445..10407
                     /codon_start=1
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     polyA_signal    10411..10459
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   12198..12260
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(12290..12310)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(12290..12308)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(12329..12345)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12329..12345)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12342..12364)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12353..12369
                     /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(12377..12383,12384..12401,12402..12407))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12422..12443
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12560..12577)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12731..13319)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12811..12830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13490..14281,14282..14350))
                     /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     14113..14132
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14351..14455)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(14530..14549)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa
       61 gaatctgctt agggttaggc gttttgcgct gcttcgggga gatgggggag gctaactgag
      121 ctctagatta accctagaaa gatagtctgc gtaaaattga cgcatgcatt cttgaaatat
      181 tgctctctct ttctaaatag cgcgaatccg tcgctgtgca tttaggacat ctcagtcgcc
      241 gcttggagct cccgtgaggc gtgcttgtca atgcggtaag tgtcactgat tttgaactat
      301 aacgaccgcg tgagtcaaaa tgacgcatga ttatctttta cgtgactttt aagatttaac
      361 tcatacgata attatattgt tatttcatgt tctacttacg tgataactta ttatatatat
      421 attttcttgt tatagatatc acaaatgctt gtccttcctt cctgtaacac agccattaaa
      481 ccaggagcat cgcccttccc cggccctcag gtaagaggac caaataccgt agccgtttcc
      541 aatttcagtc ctttagcgcc acctggtgct aactactcta tcacgctttt atccaataac
      601 tacctttgta aatttccttt caaaagttct ggccgggcgc ggtggctcac gcttgtaatc
      661 ccagcacttt gtgaggggtc aggagttcga gacgatcaac tagaatgaat gtcgagaccc
      721 atagagccca ccgcatcccc agcatgcctg ctattgtctt cccaatcctc ccccttgctg
      781 tcctgcccca ccccaccccc cagaatagaa tgacacctac tcagacaatg cgatgcaatt
      841 tcctcatttt attaggaaag gacagtggga gtggcacctt ccagggtcaa ggaaggcacg
      901 ggggaggggc aaacaacaga tggctggcaa ctagaaggca cagtcgaggc tgatcagcgg
      961 gtttaaactc aaaacagaga tgtgtcgaag atggacagtc ctgtgctgat atgcatggca
     1021 tgcaagaggc actcgtcgtt caggaaggta tccagaatct cgttcaattc cggggtcagg
     1081 ggtgagtcca ggttcagatc ctcggtcatg gactcaagtg tggttgtcag ctcatccaga
     1141 tggccccttg ggggcggatg ggaaaggtcc atttggccac agattgcagc ctcttccttc
     1201 tggggaatca cagtatcggc catctcccga agggctttga cagcctggct cgtctcttca
     1261 tcgggatcct ccaacaggtg actggcctcg ggagtcactg cgggcgctgg atccagtggc
     1321 tgagggactg gtgccggggt cagtgacccg actggctcat gtactggacc ggttggtgtt
     1381 ggtgcgaggc tggcggggag tgggcggttg gcccatggac ttcctggagg atgaaatggc
     1441 cggattcgtt ttggctggca cacctcgcgg ccctcaaaca cgtcactaat agcggagccg
     1501 gcctcaggct tcggcaaaaa catcccttcc ctggaatccc ggctgccgct gccagagccc
     1561 agcaaggctg agaaatccat atcggcgata gagctgaagt cctcgtcgcc agacagcagt
     1621 ccattaggca ggcctggtgc tcccagaggg gcaggagctg gatcaggagg cctctgagcg
     1681 cctgtcacga gccgggtgat ggcctcgggg tattccatca gcatgggctc ggtggtgtga
     1741 ggggccacag ggatgccctg gttcagcagc tgctggaact cgctgttgtc cacgctggcc
     1801 aggtcggtga acacggcagg atcggtgctg tttcccagca gggctcccag atcctcgtcg
     1861 tcgaactgca gctgcagcag agcttcagac agtgtgccct cgccggcctg tgtaggttta
     1921 ggggctggtg gagccacagc ctgtggaggt ccaggagcca gcacaggcac gggtgctggt
     1981 gcctgagcca gtgcagacac catggctgga gctggtgcag gagcaggagc ctgaggcagc
     2041 acctgaggag gggctggagc cagagcagag gcctgagaga tctggccgct ggggaacacc
     2101 atggtaggga actcgtcgta gttgatggtg ctcaggctgc tggtgaaggg gtaaggctgg
     2161 ggggcaggtt ttggcacgct ggcgctggat ctgctgggca cggcgattct tctaggtgga
     2221 ggtctagggt cggtggggcc gctgaagggg gacttcttca tgatgctctt gaatgtctcg
     2281 taggtccgct tccgcttttc ctcgatccgg tgccggtcgt cggtgtcggg caggtactgg
     2341 ctaccaactt tgcgtttctt tttcggagat ccggaacttc tagagttaat cagcatgtcc
     2401 aggtcgaaat catcaagggc gtcactgccg agcatgtcga ggtcaaagtc atcaagggca
     2461 tccgaaccaa gcatgtcaag gtcaaaatca tcgagggcgt cacttcccag catatccaga
     2521 tcaaaatcgt ccaatgcgtc agcccgtccg gaaccgctgg cctccggccc gccgctttcg
     2581 cactttagct gtttctccag gccacatatg attagttcca ggccgaaaag gaaggcaggt
     2641 tcggctccct gccggtcgaa cagctcaatt gcttgtttca gaagtggggg catagaatcg
     2701 gtggtaggtg tctctctttc ctcttttgct acttgatgct cctgttcctc caatacgcag
     2761 cccagtgtaa agtggcccac ggcggacaga gcgtacagtg cgttctccag ggagaagcct
     2821 tgctgacaca ggaacgcgag ctgattttcc agggtttcgt actgtttctc tgttgggcgg
     2881 gtgccgagat gcactttagc cccgtcgcga tgtgagagga gagcacagcg gtatgacttg
     2941 gcgttgttcc gcagaaagtc ttgccatgac tcgccttcca gggggcagga gtgggtatga
     3001 tgcctgtcca gcatctcgat tggcagggca tcgagcaggg cccgcttgtt cttcacgtgc
     3061 cagtacaggg taggctgctc aactcccagc ttttgagcga gtttccttgt cgtcaggcct
     3121 tcgataccga ctccattgag taattccaga gcagagttta tgactttgct cttgtccagt
     3181 ctagacatag aaccacctgg accagggttt tcttcaacat caccacaagt gaggagagaa
     3241 cctctacctt cgccggatcc gccctcccac acataaccag agggcagcaa ttcacgaatc
     3301 ccaactgccg tcggctgtcc atcactgtcc ttcactatgg ctttgatccc aggatgcaga
     3361 tcgagaagca cctgtcggca ccgtccgcag gggctcaaga tgcccctgtt ctcatttccg
     3421 atcgcgacga tacaagtcag gttgccagct gccgcagcag cagcagtgcc cagcaccacg
     3481 agttctgcac aaggtccccc agtaaaatga tatacattga caccagtgaa gatgcggccg
     3541 tcgctagaga gagctgcgct ggcgacgctg tagtcttcag agatggggat gctgttgatt
     3601 gtagccgttg ctctttcaat gagggtggat tcttcttgag acaaaggctt ggccatggtg
     3661 aattgctggg gagagaggtc ggtgattcgg tcaacgaggg agccgactgc cgacgtgcgc
     3721 tccggaggct tgcagaatgc ggaacaccgc gcgggcagga acagggccca cactaccgcc
     3781 ccacaccccg cctcccgcac cgccccttcc cggccgctgc tctcggcgcg ccctgctgag
     3841 cagccgctat tggccacagc ccatcgcggt cggcgcgctg ccattgctcc ctggcgctgt
     3901 ccgtctgcga gggtactagt gagacgtgcg gcttccgttt gtcacgtccg gcacgccgcg
     3961 aaccgcaagg aaccttcccg acttaggggc ggagcaggaa gcgtcgccgg ggggcccaca
     4021 agggtagcgg cgaagatccg ggtgacgctg cgaacggacg tgaagaatgt gcgagaccca
     4081 gggtcggcgc cgctgcgttt cccggaacca cgcccagagc agccgcgtcc ctgcgcaaac
     4141 ccagggctgc cttggaaaag gcgcaacccc aaccccgggg aggtggtccc tgcagttacg
     4201 ccaatgataa cccccgccag aaaaatctta gtagccttcc ctttttgttt tccgtgcccc
     4261 aactcggcgg attgactcgg ccccttccgg aaacacccga atcaacttct agtcaaatta
     4321 ttgttcacgc cgcaatgacc cacccctggc ccgcgtctgt ggaactgacc cctggtgtac
     4381 aggagagttc gctgctgaaa gtggtcccaa aggggtacta gtttttaagc tcccaactcc
     4441 ccctccccca gcgtctggag gattccacac cctcgcaccg caggggcgag gaagtgggcg
     4501 gagtccggtt ttggcgccag ccgctgaggc tgccaagcag aaaagccacc gctgaggaga
     4561 ctccggtcac tgtcctcgcc ccgcctcccc cttccctccc cttggggacc accgggcgcc
     4621 acgccgcgaa cggtaagtgc cgcggtcgtc ggcgcctccg ccctccccct agggccccaa
     4681 ttcccagcgg gcgcggcgcg cggcccctcc ccccgccggg cgcgcgcccg ctgccccgcc
     4741 cttcgtggcc gcccggcgtg ggcggtgcca cccctccccc cggcggcccc gcgcgcagct
     4801 cccggctccc tcccccttcg gatgtggctt gagctgtagg cgcggaggct agctggggat
     4861 ctggtgggct ctattggtag actttagtcc ctatcagtga tagagaaggt atctccagtt
     4921 tactccctat cagtgataga gacatccatg gaatcaactt cagctttatg agtaagaagc
     4981 cctataggat gtggctttcc ctatcagtga tagagaacta atgataagac cactgcactt
     5041 tagtccctat cagtgataga gatgagatct tctggtgtac aggagttttc tccctatcag
     5101 tgatagagag tctgtgtgcc atttgatccc tatcagtgat agagactgta tttccaggta
     5161 ggcctgtaca gtgggaggcc tatataagca gagctccttt agtgaaccct cagatggcca
     5221 gagataagag agaaaagaag agtaagaaga aatataagag ccaccacaag tttgtacaaa
     5281 aaagcaggct gccgccacca tggatgtatt catgaaagga ctttcaaagg ccaaggaggg
     5341 agttgtggct gctgctgaga aaaccaaaca gggtgtggca gaagcagcag gaaagacaaa
     5401 agagggtgtt ctctatgtag gctccaaaac caaggaggga gtggtgcatg gtgtggcaac
     5461 agtggctgag aagaccaaag agcaagtgac aaatgttgga ggagcagtgg tgacgggtgt
     5521 gacagcagta gcccagaaga cagtggaggg agcagggagc attgcagcag ccactggctt
     5581 tgtcaaaaag gaccagttgg gcaagaatga agaaggagcc ccacaggaag gaattctgga
     5641 agatatgcct gtggatcctg acaatgaggc ttatgaaatg ccttctgagg aagggtatca
     5701 agactacgaa cctgaagcct aagacccagc tttcttgtac aaagtggttg atctgctgga
     5761 agatggcgat tagagaggta cgaagatggc gattagagat ccctcccccc cccctaacgt
     5821 tactggccga agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac
     5881 catattgccg tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag
     5941 cattcctagg ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa
     6001 ggaagcagtt cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag
     6061 gcagcggaac cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga
     6121 tacacctgca aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag
     6181 agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc
     6241 ccattgtatg ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag
     6301 gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg
     6361 atgataatat ggccacaatg gactacaagg acgacgatga caagttcgtc aaatctgaga
     6421 ctctggagtt gaaggaggaa gaggaggtac tgatgctgct gggctcggct tccccggcct
     6481 cggcgaccct gaccccgatg tcctccagcg cggacgagga ggaggacgag gagctgcgcc
     6541 ggccgggctc cgcgcgtggg cagcgtggag cggaagccgg gcagggggtg cagggcagtc
     6601 cggcgtcggg tgccgggggt tgccggccag ggcggctgct gggcctgatg cacgagtgca
     6661 agcgtcgccc gtcgcgctca cgggccgtct cccgaggtgc caagacggcg gagacggtgc
     6721 agcgcatcaa gaagacccgc aggctcaagg ccaacaaccg cgagcgcaac cgcatgcaca
     6781 acctaaacgc cgcgctggac gcgctgcgcg aggtgctgcc caccttcccc gaggatgcca
     6841 agctcacgaa gatcgagacg ctgcgcttcg cccacaatta catctgggcg ctcaccgaga
     6901 ctctgcgcct ggcggaccac tgcgccggcg ccggtggcct ccagggggcg ctcttcacgg
     6961 aggcggtgct cctgagcccg ggagctgcgc tcggcgccag cggggacagc ccttctccac
     7021 cttcctcctg gagctgcacc aacagcccgg cgtcatcctc caactccacg tccccataca
     7081 gctgcacttt atcgcccgct agccccgggt cagacgtgga ctactggcag cccccacctc
     7141 cggagaagca tcgttatgcg cctcacctgc ccctcgccag ggactgtatc tctagagagg
     7201 gcaggggaag tcttctaaca tgcggggacg tggaggaaaa tcccggccca ctcgagatga
     7261 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca
     7321 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc
     7381 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     7441 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     7501 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     7561 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     7621 cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg
     7681 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     7741 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     7801 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgaaatc
     7861 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     7921 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     7981 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
     8041 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
     8101 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
     8161 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     8221 gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct
     8281 gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
     8341 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc
     8401 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctcga gagatctacg
     8461 ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt gccactccag
     8521 tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac taggtgtcct
     8581 tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt gggaagacaa
     8641 cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca caatcttggc
     8701 tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcagcct cccgagttgt
     8761 tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg tagagacggg
     8821 gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc tacccacctt
     8881 ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct gtccttgtac
     8941 cttaattaag ttaactgcta gactgcagag ggccctgcgt atgagtgcaa gtgggtttta
     9001 ggaccaggat gaggcggggt gggggtgcct acctgacgac cgaccccgac ccactggaca
     9061 agcacccaac ccccattccc caaattgcgc atcccctatc agagaggggg aggggaaaca
     9121 ggatgcggcg aggcgcgtgc gcactgccag cttcagcacc gcggacagtg ccttcgcccc
     9181 cgcctggcgg cgcgcgccac cgccgcctca gcactgaagg cgcgctgacg tcactcgccg
     9241 gtcccccgca aactcccctt cccggccacc ttggtcgcgt ccgcgccgcc gccggcccag
     9301 ccggaccgca ccacgcgagg cgcgagatag gggggcacgg gcgcgaccat ctgcgctgcg
     9361 gcgccggcga ctcagcgctg cctcagtctg cggtgggcag cggaggagtc gtgtcgtgcc
     9421 tgagagcgca gtcgagccgc caccatgaag gacaacaccg tgcccctgaa gctgatcgcc
     9481 ctgctggcca acggcgagtt ccactctggc gagcagctgg gagagaccct gggaatgagc
     9541 agagccgcca tcaacaagca catccagaca ctgagagact ggggagtgga cgtgttcacc
     9601 gtgcctggca agggctacag cctgcctgag cctatccagc tgctgaacgc caagcagatc
     9661 ctgggacagc tggatggcgg aagcgtggcc gtgctgcctg tgatcgactc caccaatcag
     9721 tacctgctgg acagaatcgg agagctgaag tccggcgacg cctgcatcgc cgagtaccag
     9781 caggctggca gaggaagaag aggacggaag tggttcagcc cattcggagc caacctgtac
     9841 ctgtccatgt tctggagact ggagcaggga cctgctgctg ccatcggact gagtctggtg
     9901 atcggaatcg tgatggccga ggtgctgaga aagctgggag ccgacaaggt gagagtgaag
     9961 tggcctaatg acctgtacct ccaggaccgc aagctggctg gcatcctggt ggagctgaca
    10021 ggcaagacag gcgatgccgc tcagatcgtg atcggagccg gaatcaacat ggccatgaga
    10081 agagtggagg agagcgtggt gaaccagggc tggatcaccc tgcaggaggc tggcatcaac
    10141 ctggaccgga acaccctggc cgccatgctg atcagagagc tgagagccgc tctggagctg
    10201 ttcgagcagg agggactggc tccttacctg agcagatggg agaagctgga caacttcatc
    10261 aacagacctg tgaagctgat catcggcgac aaggaaatct tcggcatctc cagaggaatc
    10321 gacaagcagg gagctctgct gctggagcag gacggaatca tcaagccctg gatgggcgga
    10381 gaaatctccc tgagaagcgc agagaagtaa cggcaataaa aagacagaat aaaacgcacg
    10441 ggtgttgggt cgtttgttcg cggccgcgag ctcacgggga cagccccccc cccaaagccc
    10501 ccagggatgt aattacgtcc ctcccccgct agggggcagc agcgagccgc ccggggctcc
    10561 gctccggtcc ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga cagcccgggc
    10621 acggggaagg tggcacggga tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg
    10681 cagacacctg gggggatacg gggaaaaagc tttaggctga aagagagatt tagaatgaca
    10741 gaatcataga acggcctggg ttgcaaagga gcacagtgct catccagatc caaccccctg
    10801 ctatgtgcag ggtcatcaac cagcagccca ggctgcccag agccacatcc agcctggcct
    10861 tgaatgcctg cagggatggg gcatccacag cctccttggg caacctgttc agtgcgtcac
    10921 caccctctgg gggaaaaact gcctcctcat atccaaccca aacctcccct gtctcagtgt
    10981 aaagccattc ccccttgtcc tatcaagggg gagtttgctg tgacattgtt ggtctggggt
    11041 gacacatgtt tgccaattca gtgcatcacg gagaggcaga tcttggggat aaggaagtgc
    11101 aggacagcat ggacgtggga catgcaggtg ttgagggctc tgggacactc tccaagtcac
    11161 agcgttcaga acagccttaa ggataagaag ataggataga aggacaaaga gcaagttaaa
    11221 acccagcatg gagaggagca caaaaaggcc acagacactg ctggtccctg tgtctgagcc
    11281 tgcatgtttg atggtgtctg gatgcaagca gaaggggtgg aagagcttgc ctggagagat
    11341 acagctgggt cagtaggact gggacaggca gctggagaat tgccatgtag atgttcatac
    11401 aatcgtcaaa tcatgaaggc tggaaaagcc ctccaagatc cccaagacca accccaaccc
    11461 acccaccgtg cccactggcc atgtccctca gtgccacatc cccacagttc ttcatcacct
    11521 ccagggacgg tgaccccccc acctccgtgg gcagctgtgc cactgcagca ccgctctttg
    11581 gagaaggtaa atcttgctaa atccagcccg accctcccct ggcacaacgt aaggccatta
    11641 tctctcatcc aactccagga cggagtcagt gagaatattg tttaaacatt ataggcgcgc
    11701 cattgctggg attacaggcg tgaaccactg ctcccttccc tgtcctttta aaagttttgt
    11761 tactttatag aagaaatttt gagtttttgt ttttttttaa taaataaata aacataaata
    11821 aattgtttgt tgaattywky akyasyryky wwkkgwgaag gtaaatcttg ctaaatccag
    11881 cccgaccctc ccctggcaca acgtaaggcc attatctctc atccaactcc aggacggagt
    11941 cagtgagaat attgtttaaa cattataggc gcgccattgc tgggattaca ggcgtgaacc
    12001 actgctccct tccctgtcct tttaaaagtt ttgttacttt atagaagaaa ttttgagttt
    12061 ttgttttttt ttaataaata aataaacata aataaattgt ttgttgaatt tattattagt
    12121 atgtaagtgt aaatataata aaacttaata tctattcaaa ttaataaata aacctcgata
    12181 tacagaccga taaaacacat gcgtcaattt tacgcatgat tatctttaac gtacgtcaca
    12241 atatgattat ctttctaggg ttaagcgatc gctaatccag cttttgttcc ctttagtgag
    12301 ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
    12361 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
    12421 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
    12481 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
    12541 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
    12601 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
    12661 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
    12721 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
    12781 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
    12841 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
    12901 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
    12961 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
    13021 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
    13081 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
    13141 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
    13201 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
    13261 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
    13321 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
    13381 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
    13441 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
    13501 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
    13561 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
    13621 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
    13681 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
    13741 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
    13801 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
    13861 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
    13921 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
    13981 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
    14041 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
    14101 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
    14161 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
    14221 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
    14281 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
    14341 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
    14401 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
    14461 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
    14521 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
    14581 tgtgttggag gtcgctga
//
