LOCUS       sequence-457517-       15305 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Doxycycline-inducible expression of NLS-mutated TDP-43 fused to 
            V5-tag and APEX for proximity labeling, containing BFP in the 
            backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15305)
  TITLE     Organellomics: AI-driven deep organellar phenotyping of perturbed 
            human neurons
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 15305)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: PiggyBac-TDP-43deltaNLS-V5-APEX
FEATURES             Location/Qualifiers
     source          1..15305
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(262..1053,1054..1122))
                     /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     885..904
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1123..1227)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1253..1708)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1390..1411)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1602..1621
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1835..1857
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1849..1866
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1850..1866
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   1996..2058
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        3163..3540
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    3170..3188
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3206..3224
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3242..3260
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3278..3296
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3314..3332
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3350..3368
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3386..3404
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     3449..3473
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4798..4839
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4840..5586
                     /codon_start=1
                     /product="soybean ascorbate peroxidase, engineered to serve
                     as a monomeric tag for electron microscopy and live-cell 
                     proteomics (Lam et al., 2015)"
                     /label=APEX2
                     /note="more active than the original APEX due to the A134P 
                     mutation"
                     /translation="GKSYPTVSADYQDAVEKAKKKLRGFIAEKRCAPLMLRLAFHSAGT
                     FDKGTKTGGPFGTIKHPAELAHSANNGLDIAVRLLEPLKAEFPILSYADFYQLAGVVAV
                     EVTGGPKVPFHPGREDKPEPPPEGRLPDPTKGSDHLRDVFGKAMGLTDQDIVALSGGHT
                     IGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVDK
                     YAADEDAFFADYAEAHQKLSELGFADA"
     polyA_signal    5623..5744
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5660..5679)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5714..5733
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5793..5809)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5817..5833
                     /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(5841..5847,5848..5865,5866..5871))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5886..5907
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     6333..6352
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    6351..6406
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     6452..6471
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(6562..7308)
                     /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(7340..7359)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          7367..8383
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     complement(7421..7443)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     enhancer        8663..9042
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     polyA_signal    9043..9091
                     /note="synthetic polyadenylation signal"
     misc_feature    9105..9196
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     9145..9164
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     protein_bind    complement(10408..10441)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        10442..10653
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          10654..11592
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     11549..11569
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             11615..12211
                     /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(11615..11634)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     12111..12131
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             12221..12274
                     /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             12281..12307
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             12323..12349
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     regulatory      12371..12380
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             12377..13087
                     /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    13107..13331
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(13332..13365)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     repeat_region   13881..13915
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(14406..14422)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    14430..14446
                     /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(14454..14460,14461..14478,14479..14484))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    14499..14520
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(join(14808..15305,1..91))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(14888..14907)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
       61 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
      121 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
      181 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
      241 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
      301 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
      361 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
      421 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
      481 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
      541 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
      601 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
      661 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
      721 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
      781 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
      841 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
      901 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
      961 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     1021 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     1081 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     1141 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     1201 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta
     1261 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     1321 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     1381 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     1441 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     1501 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     1561 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     1621 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     1681 gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg caactgttgg
     1741 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
     1801 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
     1861 gccagtgagc gcgcctcgtt cattcacgtt tttgaacccg tggaggacgg gcagactcgc
     1921 ggtgcaaatg tgttttacag cgtgatggag cagatgaaga tgctcgacac gctgcagaac
     1981 acgcagctag attaacccta gaaagataat catattgtga cgtacgttaa agataatcat
     2041 gtgtaaaatt gacgcatgtg ttttatcggt ctgtatatcg aggtttattt attaatttga
     2101 atagatatta agttttatta tatttacact tacatactaa taataaattc aacaaacaat
     2161 ttatttatgt ttatttattt attaaaaaaa acaaaaactc aaaatttctt ctataaagta
     2221 acaaaacttt tatgagggac agcccccccc caaagccccc agggatgtaa ttacgtccct
     2281 cccccgctag ggggcagcag cgagccgccc ggggctccgc tccggtccgg cgctcccccc
     2341 gcatccccga gccggcagcg tgcggggaca gcccgggcac ggggaaggtg gcacgggatc
     2401 gctttcctct gaacgcttct cgctgctctt tgagcctgca gacacctggg gggatacggg
     2461 gaaaaggcct ccacggcctt aattaaggga ggtggtccct gcagttacgc caatgataac
     2521 ccccgccaga aaaatcttag tagccttccc tttttgtttt ccgtgcccca actcggcgga
     2581 ttgactcggc cccttccgga aacacccgaa tcaacttcta gtcaaattat tgttcacgcc
     2641 gcaatgaccc acccctggcc cgcgtctgtg gaactgaccc ctggtgtaca ggagagttcg
     2701 ctgctgaaag tggtcccaaa ggggtactag tttttaagct cccaactccc cctcccccag
     2761 cgtctggagg attccacacc ctcgcaccgc aggggcgagg aagtgggcgg agtccggttt
     2821 tggcgccagc cgctgaggct gccaagcaga aaagccaccg ctgaggagac tccggtcact
     2881 gtcctcgccc cgcctccccc ttccctcccc ttggggacca ccgggcgcca cgccgcgaac
     2941 ggtaagtgcc gcggtcgtcg gcgcctccgc cctcccccta gggccccaat tcccagcggg
     3001 cgcggcgcgc ggcccctccc cccgccgggc gcgcgcccgc tgccccgccc ttcgtggccg
     3061 cccggcgtgg gcggtgccac ccctcccccc ggcggccccg cgcgcagctc ccggctccct
     3121 cccccttcgg atgtggcttg agctgtaggc gcggagggta ccagtttact ccctatcagt
     3181 gatagagaac gtatgaagag tttactccct atcagtgata gagaacgtat gcagacttta
     3241 ctccctatca gtgatagaga acgtataagg agtttactcc ctatcagtga tagagaacgt
     3301 atgaccagtt tactccctat cagtgataga gaacgtatct acagtttact ccctatcagt
     3361 gatagagaac gtatatccag tttactccct atcagtgata gagaacgtat aagctttagg
     3421 cgtgtacggt gggcgcctat aaaagcagag ctcgtttagt gaaccgtcag atcgcctgga
     3481 gcaattccac aacacttttg tcttatacca actttccgta ccacttccta ccctcgtaaa
     3541 ggatccgccg ccaccatgtc tgaatatatt cgggtaaccg aagatgagaa cgatgagccc
     3601 attgaaatac catcggaaga cgatgggacg gtgctgctct ccacggttac agcccagttt
     3661 ccaggggcgt gtgggcttcg ctacaggaat ccagtgtctc agtgtatgag aggtgtccgg
     3721 ctggtagaag gaattctgca tgccccagat gctggctggg gaaatctggt gtatgttgtc
     3781 aactatccaa aagataacgc agcagcaatg gatgagacag atgcttcatc agcagtgaaa
     3841 gtgaaaagag cagtccagaa aacatccgat ttaatagtgt tgggtctccc atggaaaaca
     3901 accgaacagg acctgaaaga gtattttagt acctttggag aagttcttat ggtgcaggtc
     3961 aagaaagatc ttaagactgg tcattcaaag gggtttggct ttgttcgttt tacggaatat
     4021 gaaacacaag tgaaagtaat gtcacagcga catatgatag atggacgatg gtgtgactgc
     4081 aaacttccta attctaagca aagccaagat gagcctttga gaagcagaaa agtgtttgtg
     4141 gggcgctgta cagaggacat gactgaggat gagctgcggg agttcttctc tcagtacggg
     4201 gatgtgatgg atgtcttcat ccccaagcca ttcagggcct ttgcctttgt tacatttgca
     4261 gatgatcaga ttgcgcagtc tctttgtgga gaggacttga tcattaaagg aatcagcgtt
     4321 catatatcca atgccgaacc taagcacaat agcaatagac agttagaaag aagtggaaga
     4381 tttggtggta atccaggtgg ctttgggaat cagggtggat ttggtaatag cagagggggt
     4441 ggagctggtt tgggaaacaa tcaaggtagt aatatgggtg gtgggatgaa ctttggtgcg
     4501 ttcagcatta atccagccat gatggctgcc gcccaggcag cactacagag cagttggggt
     4561 atgatgggca tgttagccag ccagcagaac cagtcaggcc catcgggtaa taaccaaaac
     4621 caaggcaaca tgcagaggga gccaaaccag gccttcggtt ctggaaataa ctcttatagt
     4681 ggctctaatt ctggtgcagc aattggttgg ggatcagcat ccaatgcagg gtcgggcagt
     4741 ggttttaatg gaggctttgg ctcaagcatg gattctaagt cttctggctg gggaatgggc
     4801 aagcccatcc ccaaccccct gctgggcctg gacagcaccg gaaagtctta cccaactgtg
     4861 agtgctgatt accaggacgc cgttgagaag gcgaagaaga agctcagagg cttcatcgct
     4921 gagaagagat gcgctcctct aatgctccgt ttggcattcc actctgctgg aacctttgac
     4981 aagggcacga agaccggtgg acccttcgga accatcaagc accctgccga actggctcac
     5041 agcgctaaca acggtcttga catcgctgtt aggcttttgg agccactcaa ggcggagttc
     5101 cctattttga gctacgccga tttctaccag ttggctggcg ttgttgccgt tgaggtcacg
     5161 ggtggaccta aggttccatt ccaccctgga agagaggaca agcctgagcc accaccagag
     5221 ggtcgcttgc ccgatcccac taagggttct gaccatttga gagatgtgtt tggcaaagct
     5281 atggggctta ctgaccaaga tatcgttgct ctatctgggg gtcacactat tggagctgca
     5341 cacaaggagc gttctggatt tgagggtccc tggacctcta atcctcttat tttcgacaac
     5401 tcatacttca cggagttgtt gagtggtgag aaggaaggtc tccttcagct accttctgac
     5461 aaggctcttt tgtctgaccc tgtattccgc cctctcgttg acaaatatgc agcggacgaa
     5521 gatgccttct ttgctgatta cgctgaggct caccaaaagc tttccgagct tgggtttgct
     5581 gatgcctgat aagtttaaac tcgcgaatcg atcgcccacc ccaacttgtt tattgcagct
     5641 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     5701 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     5761 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     5821 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     5881 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     5941 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     6001 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     6061 cggcgagcgg tatcagctca ctcaaaggac ggccgcacat gtaaggaaaa ttttagggat
     6121 gttaaagaaa aaaataacac aaaacaaaat ataaaaaaaa tctaacctca agtcaaggct
     6181 tttctatgga ataaggaatg gacagcaggg ggctgtttca tatactgatg acctctttat
     6241 agccaacctt tgttcatggc agccagcata tgggcatatg ttgccaaact ctaaaccaaa
     6301 tactcattct gatgttttaa atgatttgcc ctcccatatg tccttccgag tgagagacac
     6361 aaaaaattcc aacacactat tgcaatgaaa ataaatttcc tttattagcc agaagtcaga
     6421 tgctcaaggg gcttcatgat gtccccataa tttttggcag agggaaaaag atctcagtgg
     6481 tatttgtgag ccagggcatt ggccacacca gtcaccacct tctgataggc agcctgcacc
     6541 tgaggagtga attcacgcgt tttagcctgg cagcatatcg agatcgaagt catcgagagc
     6601 gtcagcaggg agcatatcca gatcaaaatc atccagggca tcggcgggca gcatgtccag
     6661 gtcgaagtcg tccagggcgt cggtgggtcc gccgctctcg cacttcagct gcttctcgag
     6721 gccgcagatg atcagttcca ggccgaacag gaaggcgggc tcggcgccct gtctgtcgaa
     6781 cagctcgatg gcctgcttca gcaggggggg catgctgtcg gtggtgggtg tctctctttc
     6841 ctctttggcg acctggtgtt cctgttcttc cagcacgcag cccagggtga agtggcccac
     6901 ggcgctcagg gcgtacaggg cgttttccag gctgaagccc tgctggcaca ggaaggccag
     6961 ctggttttcc agtgtctcgt actgcttctc ggtgggtctg gtgcccaggt gcactttggc
     7021 gccgtcccgg tggctcagca gggcgcatct gtagctcttg gcgttgttcc gcaggaagtc
     7081 ctgccagctc tcgccttcca gagggcagct gtgggtgtgg tgccggtcca gcatctcgat
     7141 gggcagggcg tccagcaggg cccgcttgtt cttcacgtgc cagtacaggg tgggctgttc
     7201 cacgcccagt ttctgggcca gcttccgggt ggtcaggccc tcgatgccca cgccgttcag
     7261 cagttccagg gcgctgttga tcactttgct cttgtccagc cggctcatgt cgacattctt
     7321 tgccaaaatg atgagacagc acaataacca gcacgttgcc caggagctgt aggaaaaaga
     7381 agaaggcatg aacatggtta gcagaggctc tagagccgcc ggtcacacgc cagaagccga
     7441 accccgccct gccccgtccc ccccgaaggc agccgtcccc ccgcggacag ccccgaggct
     7501 ggagagggag aaggggacgg cggcgcggcg acgcacgaag gccctccccg cccatttcct
     7561 tcctgccggc gccgcaccgc ttcgccccgc gcccgctaga gggggtgcgg cggcgcctcc
     7621 cagatttcgg ctccgcacag atttgggaca aaggaagtcc ctgcgccctc tcgcacgatt
     7681 accataaaag gcaatggctg cggctcgccg cgcctcgaca gccgccggcg ctccgggggc
     7741 cgccgcgccc cycccccgag cccyccccgg cccgmggcgg ccccgccccg cccggcaccc
     7801 ccacctgccg ccaccccccg cccggcacgg cgagccccgc gccacgcccc gtacggagcc
     7861 ccgcacccga agccgggccg tgctcagcaa ctcggggagg ggggtgcagg gggggttgca
     7921 gcccgaccga cgcgcccaca ccccctgctc acccccccac gcacacaccc cgcacgcagc
     7981 ctttgttccc ctcgcagccc cccccgcacc gcggggcacc gcccccggcc gcgctcccct
     8041 cgcgcacact gcggagcgca caaagccccg cgccgcgccc gcagcgctca cagccgccgg
     8101 gcagcgcgga gccgcacgcg gcgctcccca cgcacacaca cacgcacgca ccccccgagc
     8161 cgctcccccc gcacaaaggg ccctcccgga gcccctcaag gctttcacgc agccacagaa
     8221 aagaaacaag ccgtcattaa accaagcgct aattacagcc cggaggagaa gggccgtccc
     8281 gcccgctcac ctgtgggagt aacgcggtca gtcagagccg gggcgggcgg cgcgaggcgg
     8341 cggcggagcg gggcacgggg cgaaggcagc gcgcagcgac tcccgcccgc cgcgcgcttc
     8401 gctttttata gggccgccgc cgccgccgcc tcgccataaa aggaaacttt cggagcgcgc
     8461 cgctctgatt ggctgccgcc gcmcctcycc sccycscccc sccccccccc cccccccccc
     8521 cccccccccc cggcgcgcgc cccccccccc cccccgcccc catcgctgca caaaataatt
     8581 aaaaaataaa taaatacaaa attgggggtg gggagggggg ggagatgggg agagtgaagc
     8641 agaacgtggg gctcacctcg accatggtaa tagcgatgac taatacgtag atgtactgcc
     8701 aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc catttaccgt
     8761 cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact gccaagtggg
     8821 cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt ggcgttacta
     8881 tgggaacata cgtcattatt gacgtcaatg ggcgggggtc gttgggcggt cagccaggcg
     8941 ggccatttac cgtaagttat gtaacgcgga actccatata tgggctatga actaatgacc
     9001 ccgtaattga ttactattaa taactagtca ataatcaatg tcaataaaat atctttattt
     9061 tcattacatc tgtgtgttgg ttttttgtgt gaatcgatag tactaacata cgctctccat
     9121 caaaacaaaa cgaaacaaaa caaactagca aaataggctg tccccagtgc aagtgcaggt
     9181 gccagaacat ttctctgagc tcacggggac agcccccccc caaagccccc agggatgtaa
     9241 ttacgtccct cccccgctag ggggcagcag cgagccgccc ggggctccgc tccggtccgg
     9301 cgctcccccc gcatccccga gccggcagcg tgcggggaca gcccgggcac ggggaaggtg
     9361 gcacgggatc gctttcctct gaacgcttct cgctgctctt tgagcctgca gacacctggg
     9421 gggatacggg gaaaaagctt taggctgaaa gagagattta gaatgacaga atcatagaac
     9481 ggcctgggtt gcaaaggagc acagtgctca tccagatcca accccctgct atgtgcaggg
     9541 tcatcaacca gcagcccagg ctgcccagag ccacatccag cctggccttg aatgcctgca
     9601 gggatggggc atccacagcc tccttgggca acctgttcag tgcgtcacca ccctctgggg
     9661 gaaaaactgc ctcctcatat ccaacccaaa cctcccctgt ctcagtgtaa agccattccc
     9721 ccttgtccta tcaaggggga gtttgctgtg acattgttgg tctggggtga cacatgtttg
     9781 ccaattcagt gcatcacgga gaggcagatc ttggggataa ggaagtgcag gacagcatgg
     9841 acgtgggaca tgcaggtgtt gagggctctg ggacactctc caagtcacag cgttcagaac
     9901 agccttaagg ataagaagat aggatagaag gacaaagagc aagttaaaac ccagcatgga
     9961 gaggagcaca aaaaggccac agacactgct ggtccctgtg tctgagcctg catgtttgat
    10021 ggtgtctgga tgcaagcaga aggggtggaa gagcttgcct ggagagatac agctgggtca
    10081 gtaggactgg gacaggcagc tggagaattg ccatgtagat gttcatacaa tcgtcaaatc
    10141 atgaaggctg gaaaagccct ccaagatccc caagaccaac cccaacccac ccaccgtgcc
    10201 cactggccat gtccctcagt gccacatccc cacagttctt catcacctcc agggacggtg
    10261 acccccccac ctccgtgggc agctgtgcca ctgcagcacc gctctttgga gaaggtaaat
    10321 cttgctaaat ccagcccgac cctcccctgg cacaacgtaa ggccattatc tctcatccaa
    10381 ctccaggacg gagtcagtga gaatattata acttcgtata gcatacatta tacgaagtta
    10441 tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt cggcaattga
    10501 accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt gtactggctc
    10561 cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt
    10621 ctttttcgca acgggtttgc cgccagaaca caggtaagtg ccgtgtgtgg ttcccgcggg
    10681 cctggcctct ttacgggtta tggcccttgc gtgccttgaa ttacttccac ctggctgcag
    10741 tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt gggagagttc gaggccttgc
    10801 gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct ggcctgggcg ctggggccgc
    10861 cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg ctttcgataa gtctctagcc
    10921 atttaaaatt tttgatgacc tgctgcgacg ctttttttct ggcaagatag tcttgtaaat
    10981 gcgggccaag atctgcacac tggtatttcg gtttttgggg ccgcgggcgg cgacggggcc
    11041 cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc gagcgcggcc accgagaatc
    11101 ggacgggggt agtctcaagc tggccggcct gctctggtgc ctggcctcgc gccgccgtgt
    11161 atcgccccgc cctgggcggc aaggctggcc cggtcggcac cagttgcgtg agcggaaaga
    11221 tggccgcttc ccggccctgc tgcagggagc tcaaaatgga ggacgcggcg ctcgggagag
    11281 cgggcgggtg agtcacccac acaaaggaaa agggcctttc cgtcctcagc cgtcgcttca
    11341 tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg attagttctc gcgcttttgg
    11401 agtacgtcgt ctttaggttg gggggagggg ttttatgcga tggagtttcc ccacactgag
    11461 tgggtggaga ctgaagttag gccagcttgg cacttgatgt aattctcctt ggaatttgcc
    11521 ctttttgagt ttggatcttg gttcattctc aagcctcaga cagtggttca aagttttttt
    11581 cttccatttc aggtgtcgtg atttaaatcc caccatgacc gagtacaagc ccacggtgcg
    11641 cctcgccacc cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga
    11701 ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct
    11761 gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga
    11821 cggcgccgcc gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc
    11881 cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat
    11941 ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg
    12001 agtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga
    12061 ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc
    12121 cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg
    12181 cacctggtgc atgacccgca agcccggtgc cggatcggga gagggcagag gaagtctgct
    12241 aacatgcggt gacgtcgagg agaatcctgg cccaccggtc cctgccgcta aacgggtcaa
    12301 actcgacggc ggttctggag gtcctgctgc aaagcgcgtt aagttggatg gagggtcagg
    12361 gggagccacc gccaccatgg tgtctaaggg cgaagagctg attaaggaga acatgcacat
    12421 gaagctgtac atggagggca ccgtggacaa ccatcacttc aagtgcacat ccgagggcga
    12481 aggcaagccc tacgagggca cccagaccat gagaatcaag gtggtcgagg gcggccctct
    12541 ccccttcgcc ttcgacatcc tggctactag cttcctctac ggcagcaaga ccttcatcaa
    12601 ccacacccag ggcatccccg acttcttcaa gcagtccttc cctgagggct tcacatggga
    12661 gagagtcacc acatacgaag acgggggcgt gctgaccgct acccaggaca ccagcctcca
    12721 ggacggctgc ctcatctaca acgtcaagat cagaggggtg aacttcacat ccaacggccc
    12781 tgtgatgcag aagaaaacac tcggctggga ggccttcacc gagacgctgt accccgctga
    12841 cggcggcctg gaaggcagaa acgacatggc cctgaagctc gtgggcggga gccatctgat
    12901 cgcaaacgcc aagaccacat atagatccaa gaaacccgct aagaacctca agatgcctgg
    12961 cgtctactat gtggactaca gactggaaag aatcaaggag gccaacaacg agacctacgt
    13021 cgagcagcac gaggtggcag tggccagata ctgcgacctc cctagcaaac tggggcacaa
    13081 gcttaattag gcggccgcta agaattctgt gccttctagt tgccagccat ctgttgtttg
    13141 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
    13201 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
    13261 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
    13321 gggctctatg gataacttcg tatagcatac attatacgaa gttatttggt atggcttttt
    13381 ccccgtatcc ccccaggtgt ctgcaggctc aaagagcagc gagaagcgtt cagaggaaag
    13441 cgatcccgtg ccaccttccc cgtgcccggg ctgtccccgc acgctgccgg ctcggggatg
    13501 cggggggagc gccggaccgg agcggagccc cgggcggctc gctgctgccc cctagcgggg
    13561 gagggacgta attacatccc tgggggcttt gggggggggc tgtccctgat tctataacaa
    13621 gaaaatatat atataataag ttatcacgta agtagaacat gaaataacaa tataattatc
    13681 gtatgagtta aatcttaaaa gtcacgtaaa agataatcat gcgtcatttt gactcacgcg
    13741 gtcgttatag ttcaaaatca gtgacactta ccgcattgac aagcacgcct cacgggagct
    13801 ccaagcggcg actgagatgt cctaaatgca cagcgacgga ttcgcgctat ttagaaagag
    13861 agagcaatat ttcaagaatg catgcgtcaa ttttacgcag actatctttc tagggttaat
    13921 ctagctgcat caggatcata tcgtcgggtc ttttttccgg ctcagtcatc gcccaagctg
    13981 gcgctatctg ggcatcgggg aggaagaagc ccgtgccttt tcccgcgagg ttgaagcggc
    14041 atggaaagag tttgccgagg atgactgctg ctgcattgac gttgagcgaa aacgcacgtt
    14101 taccatgatg attcgggaag gtgtggccat gcacgccttt aacggtgaac tgttcgttca
    14161 ggccacctgg gataccagtt cgtcgcggct tttccggaca cagttccgga tggtcagccc
    14221 gaagcgcatc agcaacccga acaataccgg cgacagccgg aactgccgtg ccggtgtgca
    14281 gattaatgac agcggtgcgg cgctgggata ttacgtcagc gaggacgggt atcctggctg
    14341 gatgccgcag aaatggacat ggataccccg tgagttaccc ggcgggcgcg cttggcgtaa
    14401 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
    14461 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
    14521 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
    14581 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
    14641 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
    14701 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
    14761 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
    14821 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
    14881 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
    14941 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
    15001 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
    15061 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
    15121 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
    15181 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
    15241 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
    15301 gttgg
//
