LOCUS       sequence-462274-       14102 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Doxycycline-inducible expression of nuclear export signal (NES) 
            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 14102)
  TITLE     Organellomics: AI-driven deep organellar phenotyping of perturbed 
            human neurons
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 14102)
  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            "Unpublished"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-APEX-V5-NES
FEATURES             Location/Qualifiers
     source          1..14102
                     /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"
     regulatory      3550..3559
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3556..4305
                     /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="MGKSYPTVSAGYQDAVEKAKKKLRGFIAEKRCAPLMLRLAFHSAG
                     TFDKGTKTGGPFGTIKHPAELAHSANNGLDIAVRLLEPLKAEFPILSYADFYQLAGVVA
                     VEVTGGPKVPFHPGREDKPEPPPEGRLPDPTKGSDHLRDVFGKAMGLTDQDIVALSGGH
                     TIGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVD
                     KYAADEDAFFADYAEAHQKLSELGFADA"
     CDS             4306..4347
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4354..4380
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     polyA_signal    4420..4541
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4457..4476)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4511..4530
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4590..4606)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4614..4630
                     /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(4638..4644,4645..4662,4663..4668))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4683..4704
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     5130..5149
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    5148..5203
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     5249..5268
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(5359..6105)
                     /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(6137..6156)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          6164..7180
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     complement(6218..6240)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     enhancer        7460..7839
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     polyA_signal    7840..7888
                     /note="synthetic polyadenylation signal"
     misc_feature    7902..7993
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     7942..7961
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     protein_bind    complement(9205..9238)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        9239..9450
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          9451..10389
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     10346..10366
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             10412..11008
                     /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(10412..10431)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     10908..10928
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             11018..11071
                     /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             11078..11104
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             11120..11146
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     regulatory      11168..11177
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             11174..11884
                     /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    11904..12128
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(12129..12162)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     repeat_region   12678..12712
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(13203..13219)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    13227..13243
                     /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(13251..13257,13258..13275,13276..13281))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    13296..13317
                     /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(13605..14102,1..91))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(13685..13704)
                     /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 ccaccatggg aaagtcttac ccaactgtga gtgctggtta ccaggacgcc
     3601 gttgagaagg cgaagaagaa gctcagaggc ttcatcgctg agaagagatg cgctcctcta
     3661 atgctccgtt tggcattcca ctctgctgga acctttgaca agggcacgaa gaccggtgga
     3721 cccttcggaa ccatcaagca ccctgccgaa ctggctcaca gcgctaacaa cggtcttgac
     3781 atcgctgtta ggcttttgga gccactcaag gcggagttcc ctattttgag ctacgccgat
     3841 ttctaccagt tggctggcgt tgttgccgtt gaggtcacgg gtggacctaa ggttccattc
     3901 caccctggaa gagaggacaa gcctgagcca ccaccagagg gtcgcttgcc cgatcccact
     3961 aagggttctg accatttgag agatgtgttt ggcaaagcta tggggcttac tgaccaagat
     4021 atcgttgctc tatctggggg tcacactatt ggagctgcac acaaggagcg ttctggattt
     4081 gagggtccct ggacctctaa tcctcttatt ttcgacaact catacttcac ggagttgttg
     4141 agtggtgaga aggaaggtct ccttcagcta ccttctgaca aggctctttt gtctgaccct
     4201 gtattccgcc ctctcgttga caaatatgca gcggacgaag atgccttctt tgctgattac
     4261 gctgaggctc accaaaagct ttccgagctt gggtttgctg atgccggcaa gcccatcccc
     4321 aaccccctgc tgggcctgga cagcaccctg cagctgcctc ccctggagcg cctgaccctg
     4381 gactaataag tttaaactcg cgaatcgatc gcccacccca acttgtttat tgcagcttat
     4441 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4501 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     4561 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4621 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4681 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4741 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4801 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4861 cgagcggtat cagctcactc aaaggacggc cgcacatgta aggaaaattt tagggatgtt
     4921 aaagaaaaaa ataacacaaa acaaaatata aaaaaaatct aacctcaagt caaggctttt
     4981 ctatggaata aggaatggac agcagggggc tgtttcatat actgatgacc tctttatagc
     5041 caacctttgt tcatggcagc cagcatatgg gcatatgttg ccaaactcta aaccaaatac
     5101 tcattctgat gttttaaatg atttgccctc ccatatgtcc ttccgagtga gagacacaaa
     5161 aaattccaac acactattgc aatgaaaata aatttccttt attagccaga agtcagatgc
     5221 tcaaggggct tcatgatgtc cccataattt ttggcagagg gaaaaagatc tcagtggtat
     5281 ttgtgagcca gggcattggc cacaccagtc accaccttct gataggcagc ctgcacctga
     5341 ggagtgaatt cacgcgtttt agcctggcag catatcgaga tcgaagtcat cgagagcgtc
     5401 agcagggagc atatccagat caaaatcatc cagggcatcg gcgggcagca tgtccaggtc
     5461 gaagtcgtcc agggcgtcgg tgggtccgcc gctctcgcac ttcagctgct tctcgaggcc
     5521 gcagatgatc agttccaggc cgaacaggaa ggcgggctcg gcgccctgtc tgtcgaacag
     5581 ctcgatggcc tgcttcagca gggggggcat gctgtcggtg gtgggtgtct ctctttcctc
     5641 tttggcgacc tggtgttcct gttcttccag cacgcagccc agggtgaagt ggcccacggc
     5701 gctcagggcg tacagggcgt tttccaggct gaagccctgc tggcacagga aggccagctg
     5761 gttttccagt gtctcgtact gcttctcggt gggtctggtg cccaggtgca ctttggcgcc
     5821 gtcccggtgg ctcagcaggg cgcatctgta gctcttggcg ttgttccgca ggaagtcctg
     5881 ccagctctcg ccttccagag ggcagctgtg ggtgtggtgc cggtccagca tctcgatggg
     5941 cagggcgtcc agcagggccc gcttgttctt cacgtgccag tacagggtgg gctgttccac
     6001 gcccagtttc tgggccagct tccgggtggt caggccctcg atgcccacgc cgttcagcag
     6061 ttccagggcg ctgttgatca ctttgctctt gtccagccgg ctcatgtcga cattctttgc
     6121 caaaatgatg agacagcaca ataaccagca cgttgcccag gagctgtagg aaaaagaaga
     6181 aggcatgaac atggttagca gaggctctag agccgccggt cacacgccag aagccgaacc
     6241 ccgccctgcc ccgtcccccc cgaaggcagc cgtccccccg cggacagccc cgaggctgga
     6301 gagggagaag gggacggcgg cgcggcgacg cacgaaggcc ctccccgccc atttccttcc
     6361 tgccggcgcc gcaccgcttc gccccgcgcc cgctagaggg ggtgcggcgg cgcctcccag
     6421 atttcggctc cgcacagatt tgggacaaag gaagtccctg cgccctctcg cacgattacc
     6481 ataaaaggca atggctgcgg ctcgccgcgc ctcgacagcc gccggcgctc cgggggccgc
     6541 cgcgccccyc ccccgagccc yccccggccc grggcggccc cgccccgccc ggcacccccm
     6601 cctgccgcca ccccccgccc ggcacggcga gccccgcgcc acgccccgta cggagccccg
     6661 cacccgaagc cgggccgtgc tcagcaactc ggggaggggg gtgcaggggg ggttgcagcc
     6721 cgaccgacgc gcccacaccc cctgctcacc cccccacgca cacaccccgc acgcagcctt
     6781 tgttcccctc gcagcccccc ccgcaccgcg gggcaccgcc cccggccgcg ctcccctcgc
     6841 gcacactgcg gagcgcacaa agccccgcgc cgcgcccgca gcgctcacag ccgccgggca
     6901 gcgcggagcc gcacgcggcg ctccccacgc acacacacac gcacgcaccc cccgagccgc
     6961 tccccccgca caaagggccc tcccggagcc cctcaaggct ttcacgcagc cacagaaaag
     7021 aaacaagccg tcattaaacc aagcgctaat tacagcccgg aggagaaggg ccgtcccgcc
     7081 cgctcacctg tgggagtaac gcggtcagtc agagccgggg cgggcggcgc gaggcggcgg
     7141 cggagcgggg cacggggcga aggcagcgcg cagcgactcc cgcccgccgc gcgcttcgct
     7201 ttttataggg ccgccgccgc cgccgcctcg ccataaaagg aaactttcgg agcgcgccgc
     7261 tctgattggc tgccgccgcm ccycyccscc ycsccccscc cccccccccs cccccccccc
     7321 ccccccccgg cgcgcgcccc cccccccccc ccgcccccat cgctgcacaa aataattaaa
     7381 aaataaataa atacaaaatt gggggtgggg agggggggga gatggggaga gtgaagcaga
     7441 acgtggggct cacctcgacc atggtaatag cgatgactaa tacgtagatg tactgccaag
     7501 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
     7561 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
     7621 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
     7681 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
     7741 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
     7801 taattgatta ctattaataa ctagtcaata atcaatgtca ataaaatatc tttattttca
     7861 ttacatctgt gtgttggttt tttgtgtgaa tcgatagtac taacatacgc tctccatcaa
     7921 aacaaaacga aacaaaacaa actagcaaaa taggctgtcc ccagtgcaag tgcaggtgcc
     7981 agaacatttc tctgagctca cggggacagc ccccccccaa agcccccagg gatgtaatta
     8041 cgtccctccc ccgctagggg gcagcagcga gccgcccggg gctccgctcc ggtccggcgc
     8101 tccccccgca tccccgagcc ggcagcgtgc ggggacagcc cgggcacggg gaaggtggca
     8161 cgggatcgct ttcctctgaa cgcttctcgc tgctctttga gcctgcagac acctgggggg
     8221 atacggggaa aaagctttag gctgaaagag agatttagaa tgacagaatc atagaacggc
     8281 ctgggttgca aaggagcaca gtgctcatcc agatccaacc ccctgctatg tgcagggtca
     8341 tcaaccagca gcccaggctg cccagagcca catccagcct ggccttgaat gcctgcaggg
     8401 atggggcatc cacagcctcc ttgggcaacc tgttcagtgc gtcaccaccc tctgggggaa
     8461 aaactgcctc ctcatatcca acccaaacct cccctgtctc agtgtaaagc cattccccct
     8521 tgtcctatca agggggagtt tgctgtgaca ttgttggtct ggggtgacac atgtttgcca
     8581 attcagtgca tcacggagag gcagatcttg gggataagga agtgcaggac agcatggacg
     8641 tgggacatgc aggtgttgag ggctctggga cactctccaa gtcacagcgt tcagaacagc
     8701 cttaaggata agaagatagg atagaaggac aaagagcaag ttaaaaccca gcatggagag
     8761 gagcacaaaa aggccacaga cactgctggt ccctgtgtct gagcctgcat gtttgatggt
     8821 gtctggatgc aagcagaagg ggtggaagag cttgcctgga gagatacagc tgggtcagta
     8881 ggactgggac aggcagctgg agaattgcca tgtagatgtt catacaatcg tcaaatcatg
     8941 aaggctggaa aagccctcca agatccccaa gaccaacccc aacccaccca ccgtgcccac
     9001 tggccatgtc cctcagtgcc acatccccac agttcttcat cacctccagg gacggtgacc
     9061 cccccacctc cgtgggcagc tgtgccactg cagcaccgct ctttggagaa ggtaaatctt
     9121 gctaaatcca gcccgaccct cccctggcac aacgtaaggc cattatctct catccaactc
     9181 caggacggag tcagtgagaa tattataact tcgtatagca tacattatac gaagttatgg
     9241 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc
     9301 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     9361 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     9421 tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct
     9481 ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg gctgcagtac
     9541 gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag gccttgcgct
     9601 taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg gggccgccgc
     9661 gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc tctagccatt
     9721 taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct tgtaaatgcg
     9781 ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga cggggcccgt
     9841 gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc gagaatcgga
     9901 cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc gccgtgtatc
     9961 gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc ggaaagatgg
    10021 ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc gggagagcgg
    10081 gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt cgcttcatgt
    10141 gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgcg cttttggagt
    10201 acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca cactgagtgg
    10261 gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga atttgccctt
    10321 tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag tttttttctt
    10381 ccatttcagg tgtcgtgatt taaatcccac catgaccgag tacaagccca cggtgcgcct
    10441 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
    10501 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
    10561 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
    10621 cgccgccgtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
    10681 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
    10741 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggagt
    10801 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
    10861 ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt
    10921 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
    10981 ctggtgcatg acccgcaagc ccggtgccgg atcgggagag ggcagaggaa gtctgctaac
    11041 atgcggtgac gtcgaggaga atcctggccc accggtccct gccgctaaac gggtcaaact
    11101 cgacggcggt tctggaggtc ctgctgcaaa gcgcgttaag ttggatggag ggtcaggggg
    11161 agccaccgcc accatggtgt ctaagggcga agagctgatt aaggagaaca tgcacatgaa
    11221 gctgtacatg gagggcaccg tggacaacca tcacttcaag tgcacatccg agggcgaagg
    11281 caagccctac gagggcaccc agaccatgag aatcaaggtg gtcgagggcg gccctctccc
    11341 cttcgccttc gacatcctgg ctactagctt cctctacggc agcaagacct tcatcaacca
    11401 cacccagggc atccccgact tcttcaagca gtccttccct gagggcttca catgggagag
    11461 agtcaccaca tacgaagacg ggggcgtgct gaccgctacc caggacacca gcctccagga
    11521 cggctgcctc atctacaacg tcaagatcag aggggtgaac ttcacatcca acggccctgt
    11581 gatgcagaag aaaacactcg gctgggaggc cttcaccgag acgctgtacc ccgctgacgg
    11641 cggcctggaa ggcagaaacg acatggccct gaagctcgtg ggcgggagcc atctgatcgc
    11701 aaacgccaag accacatata gatccaagaa acccgctaag aacctcaaga tgcctggcgt
    11761 ctactatgtg gactacagac tggaaagaat caaggaggcc aacaacgaga cctacgtcga
    11821 gcagcacgag gtggcagtgg ccagatactg cgacctccct agcaaactgg ggcacaagct
    11881 taattaggcg gccgctaaga attctgtgcc ttctagttgc cagccatctg ttgtttgccc
    11941 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
    12001 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
    12061 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
    12121 ctctatggat aacttcgtat agcatacatt atacgaagtt atttggtatg gctttttccc
    12181 cgtatccccc caggtgtctg caggctcaaa gagcagcgag aagcgttcag aggaaagcga
    12241 tcccgtgcca ccttccccgt gcccgggctg tccccgcacg ctgccggctc ggggatgcgg
    12301 ggggagcgcc ggaccggagc ggagccccgg gcggctcgct gctgccccct agcgggggag
    12361 ggacgtaatt acatccctgg gggctttggg ggggggctgt ccctgattct ataacaagaa
    12421 aatatatata taataagtta tcacgtaagt agaacatgaa ataacaatat aattatcgta
    12481 tgagttaaat cttaaaagtc acgtaaaaga taatcatgcg tcattttgac tcacgcggtc
    12541 gttatagttc aaaatcagtg acacttaccg cattgacaag cacgcctcac gggagctcca
    12601 agcggcgact gagatgtcct aaatgcacag cgacggattc gcgctattta gaaagagaga
    12661 gcaatatttc aagaatgcat gcgtcaattt tacgcagact atctttctag ggttaatcta
    12721 gctgcatcag gatcatatcg tcgggtcttt tttccggctc agtcatcgcc caagctggcg
    12781 ctatctgggc atcggggagg aagaagcccg tgccttttcc cgcgaggttg aagcggcatg
    12841 gaaagagttt gccgaggatg actgctgctg cattgacgtt gagcgaaaac gcacgtttac
    12901 catgatgatt cgggaaggtg tggccatgca cgcctttaac ggtgaactgt tcgttcaggc
    12961 cacctgggat accagttcgt cgcggctttt ccggacacag ttccggatgg tcagcccgaa
    13021 gcgcatcagc aacccgaaca ataccggcga cagccggaac tgccgtgccg gtgtgcagat
    13081 taatgacagc ggtgcggcgc tgggatatta cgtcagcgag gacgggtatc ctggctggat
    13141 gccgcagaaa tggacatgga taccccgtga gttacccggc gggcgcgctt ggcgtaatca
    13201 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
    13261 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
    13321 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
    13381 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
    13441 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
    13501 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
    13561 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
    13621 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
    13681 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
    13741 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
    13801 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
    13861 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
    13921 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
    13981 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
    14041 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
    14101 gg
//
