LOCUS       sequence-468745-       10054 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For production of Extracellular Vesicles (EVs), with the Insulin 
            Receptor Strep-tag II labeled on their surface.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10054)
  AUTHORS   Gonzalez-Magaldi M, Gullapalli A, Papoulas O, Liu C, Leung AY, Guo 
            L, Brilot A, Marcotte EM, Ke Z, Leahy DJ
  TITLE     Structure and organization of full-length Epidermal Growth Factor 
            Receptor in extracellular vesicles by cryo-electron tomography.
  JOURNAL   bioRxiv [Preprint]. 2024 Nov 28:2024.11.25.625301. doi: 
            10.1101/2024.11.25.625301.
  PUBMED    39651119
REFERENCE   2  (bases 1 to 10054)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Nov 28:2024.11.25.625301. doi: 
            10.1101/2024.11.25.625301."
COMMENT     Sequence Label: pStreptagIIx2 Alfatag 3C InsR EABR
FEATURES             Location/Qualifiers
     source          1..10054
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1015..1038
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1039..1062
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1084..1122
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     CDS             1144..1167
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1189..1212
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(5645..5662)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5651..5875
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5921..6349
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6008..6027)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6218..6239
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6358..6378)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6363..6692
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6543..6678
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6759..7553
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6813..6832)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7423..7442
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7727..7848
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7764..7783)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7818..7837
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7897..7913)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7897..7913)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7910..7932)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7921..7937
                     /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(7945..7951,7952..7969,7970..7975))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7990..8011
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8128..8145)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8299..8887)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8379..8398)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9058..9849,9850..9918))
                     /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     9681..9700
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9919..10023)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gccaccatgg gcatccttcc cagccctggg atgcctgcgc tgctctccct cgtgagcctt
      961 ctctccgtgc tgctgatggg ttgcgtagct gaaggcggaa gcggcagcgg cagctggagc
     1021 cacccgcagt tcgaaaagtg gtcccacccc cagtttgaaa agtctggcag cggaggcagc
     1081 cccagcagac tggaggagga gctgaggaga aggctgaccg agcctggcag cggatctgga
     1141 tccctggaag tcctcttcca gggaccctct ggctccggat ctggaagcct tgaagtcctc
     1201 tttcagggac ccagcggctc tggcggcggg ggctcccacc tgtaccccgg agaggtgtgt
     1261 cccggcatgg atatccggaa caacctcact aggttgcatg agctggagaa ttgctctgtc
     1321 atcgaaggac acttgcagat actcttgatg ttcaaaacga ggcccgaaga tttccgagac
     1381 ctcagtttcc ccaaactcat catgatcact gattacttgc tgctcttccg ggtctatggg
     1441 ctcgagagcc tgaaggacct gttccccaac ctcacggtca tccggggatc acgactgttc
     1501 tttaactacg cgctggtcat cttcgagatg gttcacctca aggaactcgg cctctacaac
     1561 ctgatgaaca tcacccgggg ttctgtccgc atcgagaaga acaatgagct ctgttacttg
     1621 gccactatcg actggtcccg tatcctggat tccgtggagg ataattacat cgtgttgaac
     1681 aaagatgaca acgaggagtg tggagacatc tgtccgggta ccgcgaaggg caagaccaac
     1741 tgccccgcca ccgtcatcaa cgggcagttt gtcgaacgat gttggactca tagtcactgc
     1801 cagaaagttt gcccgaccat ctgtaagtca cacggctgca ccgccgaagg cctctgttgc
     1861 cacagcgagt gcctgggcaa ctgttctcag cccgacgacc ccaccaagtg cgtggcctgc
     1921 cgcaacttct acctggacgg caggtgtgtg gagacctgcc cgcccccgta ctaccacttc
     1981 caggactggc gctgtgtgaa cttcagcttc tgccaggacc tgcaccacaa atgcaagaac
     2041 tcgcggaggc agggctgcca ccagtacgtc attcacaaca acaagtgcat ccctgagtgt
     2101 ccctccgggt acacgatgaa ttccagcaac ttgctgtgca ccccatgcct gggtccctgt
     2161 cccaaggtgt gccacctcct agaaggcgag aagaccatcg actcggtgac gtctgcccag
     2221 gagctccgag gatgcaccgt catcaacggg agtctgatca tcaacattcg aggaggcaac
     2281 aatctggcag ctgagctaga agccaacctc ggcctcattg aagaaatttc agggtatcta
     2341 aaaatccgcc gatcctacgc tctggtgtca ctttccttct tccggaagtt acgtctgatt
     2401 cgaggagaga ccttggaaat tgggaactac tccttctatg ccttggacaa ccagaaccta
     2461 aggcagctct gggactggag caaacacaac ctcaccatca ctcaggggaa actcttcttc
     2521 cactataacc ccaaactctg cttgtcagaa atccacaaga tggaagaagt ttcaggaacc
     2581 aaggggcgcc aggagagaaa cgacattgcc ctgaagacca atggggacca ggcatcctgt
     2641 gaaaatgagt tacttaaatt ttcttacatt cggacatctt ttgacaagat cttgctgaga
     2701 tgggagccgt actggccccc cgacttccga gacctcttgg ggttcatgct gttctacaaa
     2761 gaggcccctt atcagaatgt gacggagttc gacgggcagg atgcatgtgg ttccaacagt
     2821 tggacggtgg tagacattga cccacccctg aggtccaacg accccaaatc acagaaccac
     2881 ccagggtggc tgatgcgggg tctcaagccc tggacccagt atgccatctt tgtgaagacc
     2941 ctggtcacct tttcggatga acgccggacc tatggggcca agagtgacat catttatgtc
     3001 cagacagatg ccaccaaccc ctctgtgccc ctggatccaa tctcagtgtc taactcatca
     3061 tcccagatta ttctgaagtg gaaaccaccc tccgacccca atggcaacat cacccactac
     3121 ctggttttct gggagaggca ggcggaagac agtgagctgt ttgagctgga ttattgcctc
     3181 aaagggctga agctgccctc gaggacctgg tctccaccat tcgagtctga agattctcag
     3241 aagcacaacc agagtgagta tgaggattcg gccggcgaat gctgctcctg tccaaagaca
     3301 gactctcaga tcctgaagga gctggaggag tcctcgttta ggaagacgtt tgaggattac
     3361 ctgcacaacg tggttttcgt ccccaggcca tctcggaaac gcaggtccct tggcgatgtt
     3421 gggaatgtga cggtggccgt gcccacggtg gcagctttcc ccaacacttc ctcgaccagc
     3481 gtgcccacga gtccggagga gcacaggcct tttgagaagg tggtgaacaa ggagtcgctg
     3541 gtcatctccg gcttgcgaca cttcacgggc tatcgcatcg agctgcaggc ttgcaaccag
     3601 gacacccctg aggaacggtg cagtgtggca gcctacgtca gtgcgaggac catgcctgaa
     3661 gccaaggctg atgacattgt tggccctgtg acgcatgaaa tctttgagaa caacgtcgtc
     3721 cacttgatgt ggcaggagcc gaaggagccc aatggtctga tcgtgctgta tgaagtgagt
     3781 tatcggcgat atggtgatga ggagctgcat ctctgcgtct cccgcaagca cttcgctctg
     3841 gaacggggct gcaggctgcg tgggctgtca ccggggaact acagcgtgcg aatccgggcc
     3901 acctcccttg cgggcaacgg ctcttggacg gaacccacct atttctacgt gacagactat
     3961 ttagacgtcc cgtcaaatat tgcaaaaatt atcatcggcc ccctcatctt tgtctttctc
     4021 ttcagtgttg tgattggaag tatttatcta ttcctgagaa agaggcagcc agatgggccg
     4081 ctgggaccgc tttacgcttc ttcaaaccct gagtatctca gtgccagtga tgtgtttcca
     4141 tgctctgtgt acgtgccgga cgagtgggag gtgtctcgag agaagatcac cctccttcga
     4201 gagctggggc agggctcctt cggcatggtg tatgagggca atgccaggga catcatcaag
     4261 ggtgaggcag agacccgcgt ggcggtgaag acggtcaacg agtcagccag tctccgagag
     4321 cggattgagt tcctcaatga ggcctcggtc atgaagggct tcacctgcca tcatgtggtg
     4381 cgcctcctgg gagtggtgtc caagggccag cccacgctgg tggtgatgga gctgatggct
     4441 cacggagacc tgaagagcta cctccgttct ctgcggccag aggctgagaa taatcctggc
     4501 cgccctcccc ctacccttca agagatgatt cagatggcgg cagagattgc tgacgggatg
     4561 gcctacctga acgccaagaa gtttgtgcat cgggacctgg cagcgagaaa ctgcatggtc
     4621 gcccatgatt ttactgtcaa aattggagac tttggaatga ccagagacat ctatgaaacg
     4681 gattactacc ggaaaggggg caagggtctg ctccctgtac ggtggatggc accggagtcc
     4741 ctgaaggatg gggtcttcac cacttcttct gacatgtggt cctttggcgt ggtcctttgg
     4801 gaaatcacca gcttggcaga acagccttac caaggcctgt ctaatgaaca ggtgttgaaa
     4861 tttgtcatgg atggagggta tctggatcaa cccgacaact gtccagagag agtcactgac
     4921 ctcatgcgca tgtgctggca attcaacccc aagatgaggc caaccttcct ggagattgtc
     4981 aacctgctca aggacgacct gcaccccagc tttccagagg tgtcgttctt ccacagcgag
     5041 gagaacaagg ctcccgagag tgaggagctg gagatggagt ttgaggacat ggagaatgtg
     5101 cccctggacc gttcctcgca ctgtcagagg gaggaggcgg ggggccggga tggagggtcc
     5161 tcgctgggtt tcaagcggag ctacgaggaa cacatccctt acacacacat gaacggaggc
     5221 aagaaaaacg ggcggattct gaccttgcct cggtccaatc cttccagcgg ccgcgctttg
     5281 cctggcaatc ccgatcacag ggaaatgggg gaaactctcc cagaagaggt tggagaatat
     5341 cggcagccat ctggtggcag tgtacctgtg tctccgggtc cgcccagtgg tcttgaaccc
     5401 actagctcct caccttacgg cggcggctcc tttaacagct caatcaacaa catccacgaa
     5461 atggagattc agctgaagga tgcactcgaa aagaatcaac aatggctcgt ttacgaccaa
     5521 caacgcgagg tgtacgtgaa aggattgctt gcaaaaattt tcgaactgga aaagaaaaca
     5581 gagactgcag cacatagtct tccttgactc gagtctagag ggcccgttta aacccgctga
     5641 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     5701 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     5761 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     5821 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     5881 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     5941 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     6001 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     6061 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     6121 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     6181 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     6241 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     6301 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     6361 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     6421 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     6481 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     6541 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     6601 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     6661 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     6721 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     6781 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     6841 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     6901 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     6961 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     7021 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     7081 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     7141 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     7201 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     7261 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     7321 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     7381 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     7441 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     7501 ctccagattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     7561 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     7621 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     7681 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     7741 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     7801 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     7861 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     7921 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     7981 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     8041 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     8101 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     8161 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     8221 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     8281 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     8341 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     8401 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     8461 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     8521 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     8581 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     8641 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     8701 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     8761 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     8821 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     8881 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     8941 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     9001 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     9061 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     9121 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     9181 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     9241 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     9301 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     9361 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     9421 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     9481 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     9541 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     9601 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     9661 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     9721 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     9781 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     9841 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     9901 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     9961 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
    10021 gcgcacattt ccccgaaaag tgccacctga cgtc
//
