LOCUS       sequence-365349-       14165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Big-IN Payload yeast assembly vector. Encodes LEU2 yeast selectable 
            marker and components for bacterial copy number induction. Contains 
            a mammalian transient (backbone) GFP-T2A-BSD selection cassette..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14165)
  AUTHORS   Brosh R, Laurent JM, Ordonez R, Huang E, Hogan MS, Hitchcock AM, 
            Mitchell LA, Pinglay S, Cadley JA, Luther RD, Truong DM, Boeke JD, 
            Maurano MT
  TITLE     A versatile platform for locus-scale genome rewriting and 
            verification.
  JOURNAL   Proc Natl Acad Sci U S A. 2021 Mar 9;118(10). pii: 2023952118. doi: 
            10.1073/pnas.2023952118.
  PUBMED    33649239
REFERENCE   2  (bases 1 to 14165)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2021-03-9"; volume: "118(10). pii: 
            2023952118. doi"; pages: " 10.1073/pnas.2023952118"
COMMENT     Sequence Label: pLM1110_LEU2
FEATURES             Location/Qualifiers
     source          1..14165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        433..1611
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          664..1602
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1559..1579
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             join(1632..1634,1635..1637,1638..2348)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1632..1634,1635..1637,1638..2348)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1677..1698)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1938..1957)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2285..2306
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2358..2411
                     /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             2412..2810
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(2838..2855)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2844..3051
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(3269..3302)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             complement(3356..4033)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             4040..4051
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    4069..4085
                     /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(4093..4099,4100..4117,4118..4123))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4138..4159
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    complement(4270..4303)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             4784..5878
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     CDS             complement(6455..7270)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(6455..7270)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     7178..7197
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(7271..7373)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     mobile_element  7401..8168
                     /mobile_element_type="insertion sequence:IS1"
                     /label=IS1
                     /note="prokaryotic transposable element"
     misc_feature    8170..8673
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     rep_origin      9524..10138
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"
     rep_origin      9615..10146
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      10214..10433
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     CDS             10524..11279
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     misc_feature    11282..11532
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             11858..13033
                     /codon_start=1
                     /gene="sopA"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopA
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     CDS             13033..14004
                     /codon_start=1
                     /gene="sopB"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopB
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
ORIGIN
        1 tctgattatt agtctgggac cacggtccca ctcgtatcgt cggtctgatt attagtctgg
       61 gaccacggtc ccactcgtat cgtcggtctg atwatyagwc tggraccacg gtcccactcg
      121 tatcgtcggt ctgattatta gtctgggacc acggtcccac tcgtatcgtc ggtctgatta
      181 ttagtctggg accacgatcc cactcgtgtt gtcggtctga ttatcggtct gggaccacgg
      241 tcccacttgt attgtcgatc agactatcag cgtgagacta cgattccatc aatgcctgtc
      301 aagggcaagt attgacatgt cgtcgtaacc tgtagaacgg agtaacctcg gtgtgcggtt
      361 gtatgcctgc ggtggattgc tgctgtgtcc tgcttatcca caacattttg cgcacggtta
      421 tgtggacggg ccggctccgg tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc
      481 gagaagttgg ggggaggggt cggcaattga accggtgcct agagaaggtg gcgcggggta
      541 aactgggaaa gtgatgtcgt gtactggctc cgcctttttc ccgagggtgg gggagaaccg
      601 tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca
      661 caggtaagtg ccgtgtgtgg ttcccgcggg cctggcctct ttacgggtta tggcccttgc
      721 gtgccttgaa ttacttccac ctggctgcag tacgtgattc ttgatcccga gcttcgggtt
      781 ggaagtgggt gggagagttc gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga
      841 gttgaggcct ggcctgggcg ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc
      901 tgtctcgctg ctttcgataa gtctctagcc atttaaaatt tttgatgacc tgctgcgacg
      961 ctttttttct ggcaagatag tcttgtaaat gcgggccaag atctgcacac tggtatttcg
     1021 gtttttgggg ccgcgggcgg cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg
     1081 cggggcctgc gagcgcggcc accgagaatc ggacgggggt agtctcaagc tggccggcct
     1141 gctctggtgc ctagtctcgc gccgccgtgt atcgccccgc cctgggcggc aaggctggcc
     1201 cggtcggcac cagttgcgtg agcggaaaga tggccgcttc ccggccctac tgcagggagc
     1261 tcaaaatgga ggacgcggcg ctcgggagag cgggcgggtg agtcacccac acaaaggaaa
     1321 agggcctttc cgtcctcagc cgtcgcttca tgtgactcca cggagtaccg ggcgctgtcc
     1381 aggcacctcg attagttctc gagcttttgg agtacgtcgt ctttaggttg gggggagggg
     1441 ttttatgcga tggagtttcc ccacactgag tgggtggaga ctgaagttag gccagcttgg
     1501 cacttgatgt aattctcctt ggaatttgcc ctttttgagt ttggatcttg gttcattctc
     1561 aagcctcaga cagtggttca aagttttttt cttccatttc aggtgtcgtg atctctagaa
     1621 gctgggtacc aatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     1681 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     1741 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     1801 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     1861 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     1921 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     1981 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     2041 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     2101 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     2161 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     2221 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     2281 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     2341 tgtacaaggg cagtggagag ggcagaggaa gtctgctaac atgcggtgac gtcgaggaga
     2401 atcctggccc aatggccaag cctttgtctc aagaagaatc caccctcatt gaaagagcaa
     2461 cggctacaat caacagcatc cccatctctg aggactacag cgtcgccagc gcagctctct
     2521 ctagcgacgg ccgcatcttc actggtgtca atgtatatca ttttactggg ggaccttgtg
     2581 cagaactcgt ggtgctgggc actgctgctg ctgcggcagc tggcaacctg acttgtatcg
     2641 tcgcgatcgg aaatgagaac aggggcatct tgagcccctg cggacggtgc cgacaggtgc
     2701 ttctcgatct gcatcctggg atcaaagcca tagtgaagga cagtgatgga cagccgacgg
     2761 cagttgggat tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa gaattctaac
     2821 tagagctcgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     2881 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     2941 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     3001 gcaggacagc aagggggagg attgggaaga gaatagcagg catgctgggg ataaggtact
     3061 accaatattt agtttctaag ccttgcgaca gacctcccac ttagattgcc acgcatagag
     3121 ctagcgagtc agcgaaaagc atgacgcgct ttcaagcgtg gcgagtatgt gaaccaaggc
     3181 ttcggacagg actatatact taggtttgat ctcgccccga gaactgtaaa cctcaacatt
     3241 tatagattat tagggataac agggtaatat aacttcgtat aggatacttt atacgaagtt
     3301 atcagtagag accgcctggc tctagtagcg atctacacta gcactatcag cgttattaag
     3361 caccggtgga gtgacgacct tcagcacgtt cgtactgttc aacgatggtg tagtcttcgt
     3421 tgtgggaggt gatgtccagt ttgatgtcgg ttttgtaagc acccggcagc tgaaccggtt
     3481 ttttagccat gtaggtggtt ttaacttcag cgtcgtagtg accaccgtct ttcagtttca
     3541 gacgcatttt gatttcacct ttcagagcac cgtcttccgg gtacatacgt tcggtggaag
     3601 cttcccaacc catggttttt ttctgcataa ccggaccgtc ggacgggaag ttggtaccac
     3661 gcagtttaac tttgtagatg aactcaccgt cttgcaggga ggagtcctgg gtaacggtaa
     3721 caacaccacc gtcttcgaag ttcataacac gttcccattt gaaaccttcc gggaaggaca
     3781 gtttcaggta gtccgggatg tcagccgggt gtttaacgta agctttggaa ccgtactgga
     3841 actgcgggga caggatgtcc caagcgaacg gcagcggacc acctttggta actttcagtt
     3901 tagcggtctg ggtaccttcg tacggacgac cttcaccttc accttcgatt tcgaactcgt
     3961 gaccgttaac ggaaccttcc atacgaactt tgaaacgcat gaactctttg ataacgtctt
     4021 cggaggaagc catctagtat ttctcctctt tctctagtat gtgtgaaatt gttatccgct
     4081 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     4141 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     4201 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgattggg
     4261 tctcatttta taacttcgta tagcatacat tatacgaagt tattagggat aacagggtaa
     4321 ttttagggta gcatcaggaa tctgaaccct cagaaagtgg ggatcccggg tatagacctt
     4381 tatctgcggt tcaagttagg cataaggctg catgctacct tgtcacacct acactgctcg
     4441 aagtaaatat gggaagcgtg cgacctggct ccaggcgttc cgcgccgcca cgtgttcgtt
     4501 aactgttgat tggtggcaca tagaggtcgc ctgacgcata tacctttttc aactgaaaaa
     4561 ttgggagaaa aaggaaaggt gagagcgccg gaaccggctt ttcatataga atagagaagc
     4621 gttcatgact aaatgcttgc atcacaatac ttgaagttga caatattatt taaggaccta
     4681 ttgttttttc caataggtgg ttagcaatcg tcttactttc taacttttct taccttttac
     4741 atttcagcaa tatatatata tatttcaagg atataccatt ctaatgtctg cccctaagaa
     4801 gatcgtcgtt ttgccaggtg accacgttgg tcaagaaatc acagccgaag ccattaaggt
     4861 tcttaaagct atttctgatg ttcgttccaa tgtcaagttc gatttcgaaa atcatttaat
     4921 tggtggtgct gctatcgatg ctacaggtgt tccacttcca gatgaggcgc tggaagcctc
     4981 caagaaggct gatgccgttt tgttaggtgc tgtgggtggt cctaaatggg gtaccggtag
     5041 tgttagacct gaacaaggtt tactaaaaat ccgtaaagaa cttcaattgt acgccaactt
     5101 aagaccatgt aactttgcat ccgactctct tttagactta tctccaatca agccacaatt
     5161 tgctaaaggt actgacttcg ttgttgtcag agaattagtg ggaggtattt actttggtaa
     5221 gagaaaggaa gacgatggtg atggtgtcgc ttgggatagt gaacaataca ccgttccaga
     5281 agtgcaaaga atcacaagaa tggccgcttt catggcccta caacatgagc caccattgcc
     5341 tatttggtcc ttggataaag ctaatgtttt ggcctcttca agattatgga gaaaaactgt
     5401 ggaggaaacc atcaagaacg aattccctac attgaaggtt caacatcaat tgattgattc
     5461 tgccgccatg atcctagtta agaacccaac ccacctaaat ggtattataa tcaccagcaa
     5521 catgtttggt gatatcatct ccgatgaagc ctccgttatc ccaggttcct tgggtttgtt
     5581 gccatctgcg tccttggcct ctttgccaga caagaacacc gcatttggtt tgtacgaacc
     5641 atgccacggt tctgctccag atttgccaaa gaataaggtc aaccctatcg ccactatctt
     5701 gtctgctgca atgatgttga aattgtcatt gaacttgcct gaagaaggta aggccattga
     5761 agatgcagtt aaaaaggttt tggatgcagg tatcagaact ggtgatttag gtggttccaa
     5821 cagtaccacc gaagtcggtg atgctgtcgc cgaagaagtt aagaaaatcc ttgcttaaaa
     5881 agattctctt tttttatgat atttgtacat aaactttata aatgaaattc ataatagaaa
     5941 cgacacgaaa ttacaaaatg gaatatgttc atagggtaga cgaaactata tacgcaatct
     6001 acatacattt atcaagaagg agaaaaagga ggatgtaaag gaatacaggt aagcaaattg
     6061 atactaatgg ctcaacgtga taaggaaaaa gaattgcact ttaacattaa tattgacaag
     6121 gaggagggca ccgcggccgc ccaactacgc agccgatcaa caacagaaac gcgacggtcg
     6181 agatgaggaa cgcacactgg agcctttatc agatggtgtt gatttcaatt actccggagc
     6241 gtgatgcgga tcaaagtaac cgcgatggaa ggtgggggac tcgaaataga tccgttctta
     6301 gccctgcgaa acatctactg tacaccacat gcaacagcca caattctcat gtttgacagc
     6361 ttatcatcga atttctgcca ttcatccgct tattatcact tattcaggcg tagcaaccag
     6421 gcgtttaagg gcaccaataa ctgccttaaa aaaattagaa aaactcatcg agcatcaaat
     6481 gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa agccgtttct
     6541 gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt
     6601 ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa
     6661 ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt
     6721 tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac
     6781 tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agccagacga aatacgcgat
     6841 cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca
     6901 gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt
     6961 ttccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga
     7021 tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat
     7081 cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat
     7141 acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat
     7201 ataaatcagc atccatgttg gaatttaatc gcggcctcga gcaagacgtt tcccgttgaa
     7261 tatggctcat tttagcttcc ttagctcctg aaaatctcga taactcaaaa aatacgtccg
     7321 gtagtgatct tatttcatta tggtgaaagt tggaacctct tacgtgccga tcaacgtccc
     7381 tgggtccttt tcatcacgtg ggtgatgctg ccaacttact gatttagtgt atgatggtgt
     7441 ttttgaggtg ctccagtggc ttctgtttct atcagctgtc cctcctgttc agctactgac
     7501 ggggtggtgc gtaacggcaa aagcaccgcc ggacatcagc gctatctctg ctctcactgc
     7561 cgtaaaacat ggcaactgca gttcacttac accgcttctc aacccggtac gcaccagaaa
     7621 atcattgata tggccatgaa tggcgttgga tgccgggcaa cagcccgcat tatgggcgtt
     7681 ggcctcaaca cgattttacg tcacttaaaa aactcaggcc gcagtcggta acctcgcgca
     7741 tacagccggg cagtgacgtc atcgtctgcg cggaaatgga cgaacagtgg ggctatgtcg
     7801 gggctaaatc gcgccagcgc tggctgtttt acgcgtatga cagtctccgg aagacggttg
     7861 ttgcgcacgt attcggtgaa cgcactatgg cgacgctggg gcgtcttatg agcctgctgt
     7921 caccctttga cgtggtgata tggatgacgg atggctggcc gctgtatgaa tcccgcctga
     7981 agggaaagct gcacgtaatc agcaagcgat atacgcagcg aattgagcgg cataacctga
     8041 atctgaggca gcacctggca cggctgggac ggaagtcgct gtcgttctca aaatcggtgg
     8101 agctgcatga caaagtcatc gggcattatc tgaacataaa acactatcaa taagttggag
     8161 tcattaccca tcacgtgcta taaaaataat tataatttaa attttttaat ataaatatat
     8221 aaattaaaaa tagaaagtaa aaaaagaaat taaagaaaaa atagtttttg ttttccgaag
     8281 atgtaaaaga ctctaggggg atcgccaaca aatactacct tttatcttgc tcttcctgct
     8341 ctcaggtatt aatgccgaat tgtttcatct tgtctgtgta gaagaccaca cacgaaaatc
     8401 ctgtgatttt acattttact tatcgttaat cgaatgtata tctatttaat ctgcttttct
     8461 tgtctaataa atatatatgt aaagtacgct ttttgttgaa attttttaaa cctttgttta
     8521 tttttttttc ttcattccgt aactcttcta ccttctttat ttactttcta aaatccaaat
     8581 acaaaacata aaaataaata aacacagagt aaattcccaa attattccat cattaaaaga
     8641 tacgaggcgc gtgtaagtta caggcaagcg atcttttcgc caaaagttgg cccagggctt
     8701 cccggtatca acagggacac caggatttat ttattctgcg aagtgatctt ccgtcacagg
     8761 tatttattcg cgataagctc atggagcggc gtaaccgtcg cacaggaagg acagagaaag
     8821 cgcggatctg ggaagtgacg gacagaacgg tcaggacctg gattggggag gcggttgccg
     8881 ccgctgctgc tgacggtgtg acgttctctg ttccggtcac accacatacg ttccgccatt
     8941 cctatgcgat gcacatgctg tatgccggta taccgctgaa agttctgcaa agcctgatgg
     9001 gacataagtc catcagttca acggaagtct acacgaaggt ttttgcgctg gatgtggctg
     9061 cccggcaccg ggtgcagttt gcgatgccgg agtctgatgc ggttgcgatg ctgaaacaat
     9121 tatcctgaga ataaatgcct tggcctttat atggaaatgt ggaactgagt ggatatgctg
     9181 tttttgtctg ttaaacagag aagctggctg ttatccactg agaagcgagc gaaacagtcg
     9241 ggaaaatctc ccattatcgt agagatccgc attattaatc tcaggagcct gtgtagcgtt
     9301 tataggaagt agtgttctgt catgatgcct gcaagcggta acgaaaacga tttgaatatg
     9361 ccttcaggaa caatagaaat cttcgtgcgg tgttacgttg aagtggagcg gattatgtca
     9421 gcaatggaca gaacaaccta atgaacacag aaccatgatg tggtctgtcc ttttacagcc
     9481 agtagtgctc gccgcagtcg agcgacaggg cgaagccctc gagctggttg ccctcgccgc
     9541 tgggctggcg gctgtctatg gccctgcaaa cgcgccagaa acgccgtcga agccgtgtgc
     9601 gagacaccgc ggccggccgc cggcgttgtg gatacctcgc ggaaaacttg gccctcactg
     9661 acagatgagg ggcggacgtt gacacttgag gggccgactc acccggcgcg gcgttgacag
     9721 atgaggggca ggctcgattt cggccggcga cgtggagctg gacagcctcg caaatcggcg
     9781 aaaacgcctg attttacgcg agtttcccac agatgatgtg gacaagcctg gggataagtg
     9841 ccctgcggta ttgacacttg aggggcgcga ctactgacag atgaggggcg cgatccttga
     9901 cacttgaggg gcagagtgct gacagatgag gggcgcacct attgacattt gaggggctgt
     9961 ccacaggcag aaaatccagc atttgcaagg gtttccgccc gtttttcggc caccgctaac
    10021 ctgtctttta acctgctttt aaaccaatat ttataaacct tgtttttaac cagggctgcg
    10081 ccctgtgcgc gtgaccgcgc acgccgaagg ggggtgcccc cccttctcga accctcccgg
    10141 tcgagtgagc gaggaagcac cagggaacag cacttatata ttctgcttac acacgatgcc
    10201 tgaaaaaact tcccttgggg ttatccactt atccacgggg atatttttat aattattttt
    10261 tttatagttt ttagatcttc ttttttagag cgccttgtag gcctttatcc atgctggttc
    10321 tagagaaggt gttgtgacaa attgcccttt cagtgtgaca aatcaccctc aaatgacagt
    10381 cctgtctgtg acaaattgcc cttaaccctg tgacaaattg ccctcagaag aagctgtttt
    10441 ttcacaaagt tatccctgct tattgactct tttttattta gtgtgacaat ctaaaaactt
    10501 gtcacacttc acatggatct gtcatggcgg aaacagcggt tatcaatcac aagaaacgta
    10561 aaaatagccc gcgaatcgtc cagtcaaacg acctcactga ggcggcatat agtctctccc
    10621 gggatcaaaa acgtatgctg tatctgttcg ttgaccagat cagaaaatct gatggcaccc
    10681 tacaggaaca tgacggtatc tgcgagatcc atgttgctaa atatgctgaa atattcggat
    10741 tgacctctgc ggaagccagt aaggatatac ggcaggcatt gaagagtttc gcggggaagg
    10801 aagtggtttt ttatcgccct gaagaggatg ccggcgatga aaaaggctat gaatcttttc
    10861 cttggtttat caaacgtgcg cacagtccat ccagagggct ttacagtgta catatcaacc
    10921 catatctcat tcccttcttt atcgggttac agaaccggtt tacgcagttt cggcttagtg
    10981 aaacaaaaga aatcaccaat ccgtatgcca tgcgtttata cgaatccctg tgtcagtatc
    11041 gtaagccgga tggctcaggc attgtctctc tgaaaatcga ctggatcata gagcgttacc
    11101 agctgcctca aagttaccag cgtatgcctg acttccgccg ccgcttcctg caggtctgtg
    11161 ttaatgagat caacagcaga actccaatgc gcctctcata cattgagaaa aagaaaggcc
    11221 gccagacgac tcatatcgta ttttccttcc gcgatatcac ttccatgacg acaggatagt
    11281 ctgagggtta tctgtcacag atttgagggt ggttcgtcac atttgttctg acctactgag
    11341 ggtaatttgt cacagttttg ctgtttcctt cagcctgcat ggattttctc atactttttg
    11401 aactgtaatt tttaaggaag ccaaatttga gggcagtttg tcacagttga tttccttctc
    11461 tttcccttcg tcatgtgacc tgatatcggg ggttagttcg tcatcattga tgagggttga
    11521 ttatcacagt ttattactct gaattggcta tccgcgtgtg tacctctacc tggagttttt
    11581 cccacggtgg atatttcttc ttgcgctgag cgtaagagct atctgacaga acagttcttc
    11641 tttgcttcct cgccagttcg ctcgctatgc tcggttacac ggctgcggcg agcgctagtg
    11701 ataataagtg actgaggtat gtgctcttct tatctccttt tgtagtgttg ctcttatttt
    11761 aaacaacttt gcggtttttt gatgactttg cgattttgtt gttgctttgc agtaaattgc
    11821 aagatttaat aaaaaaacgc aaagcaatga ttaaaggatg ttcagaatga aactcatgga
    11881 aacacttaac cagtgcataa acgctggtca tgaaatgacg aaggctatcg ccattgcaca
    11941 gtttaatgat gacagcccgg aagcgaggaa aataacccgg cgctggagaa taggtgaagc
    12001 agcggattta gttggggttt cttctcaggc tatcagagat gccgagaaag cagggcgact
    12061 accgcacccg gatatggaaa ttcgaggacg ggttgagcaa cgtgttggtt atacaattga
    12121 acaaattaat catatgcgtg atgtgtttgg tacgcgattg cgacgtgctg aagacgtatt
    12181 tccaccggtg atcggggttg ctgcccataa aggtggcgtt tacaaaacct cagtttctgt
    12241 tcatcttgct caggatctgg ctctgaaggg gctacgtgtt ttgctcgtgg aaggtaacga
    12301 cccccaggga acagcctcaa tgtatcacgg atgggtacca gatcttcata ttcatgcaga
    12361 agacactctc ctgcctttct atcttgggga aaaggacgat gtcacttatg caataaagcc
    12421 cacttgctgg ccggggcttg acattattcc ttcctgtctg gctctgcacc gtattgaaac
    12481 tgagttaatg ggcaaatttg atgaaggtaa actgcccacc gatccacacc tgatgctccg
    12541 actggccatt gaaactgttg ctcatgacta tgatgtcata gttattgaca gcgcgcctaa
    12601 cctgggtatc ggcacgatta atgtcgtatg tgctgctgat gtgctgattg ttcccacgcc
    12661 tgctgagttg tttgactaca cctccgcact gcagtttttc gatatgcttc gtgatctgct
    12721 caagaacgtt gatcttaaag ggttcgagcc tgatgtacgt attttgctta ccaaatacag
    12781 caatagcaat ggctctcagt ccccgtggat ggaggagcaa attcgggatg cctggggaag
    12841 catggttcta aaaaatgttg tacgtgaaac ggatgaagtt ggtaaaggtc agatccggat
    12901 gagaactgtt tttgaacagg ccattgatca acgctcttca actggtgcct ggagaaatgc
    12961 tctttctatt tgggaacctg tctgcaatga aattttcgat cgtctgatta aaccacgctg
    13021 ggagattaga taatgaagcg tgcgcctgtt attccaaaac atacgctcaa tactcaaccg
    13081 gttgaagata cttcgttatc gacaccagct gccccgatgg tggattcgtt aattgcgcgc
    13141 gtaggagtaa tggctcgcgg taatgccatt actttgcctg tatgtggtcg ggatgtgaag
    13201 tttactcttg aagtgctccg gggtgatagt gttgagaaga cctctcgggt atggtcaggt
    13261 aatgaacgtg accaggagct gcttactgag gacgcactgg atgatctcat cccttctttt
    13321 ctactgactg gtcaacagac accggcgttc ggtcgaagag tatctggtgt catagaaatt
    13381 gccgatggga gtcgccgtcg taaagctgct gcacttaccg aaagtgatta tcgtgttctg
    13441 gttggcgagc tggatgatga gcagatggct gcattatcca gattgggtaa cgattatcgc
    13501 ccaacaagtg cttatgaacg tggtcagcgt tatgcaagcc gattgcagaa tgaatttgct
    13561 ggaaatattt ctgcgctggc tgatgcggaa aatatttcac gtaagattat tacccgctgt
    13621 atcaacaccg ccaaattgcc taaatcagtt gttgctcttt tttctcaccc cggtgaacta
    13681 tctgcccggt caggtgatgc acttcaaaaa gcctttacag ataaagagga attacttaag
    13741 cagcaggcat ctaaccttca tgagcagaaa aaagctgggg tgatatttga agctgaagaa
    13801 gttatcactc ttttaacttc tgtgcttaaa acgtcatctg catcaagaac tagtttaagc
    13861 tcacgacatc agtttgctcc tggagcgaca gtattgtata agggcgataa aatggtgctt
    13921 aacctggaca ggtcccgtgt tccaactgag tgtatagaga aaattgaggc cattcttaag
    13981 gaacttgaaa agccagcacc ctgatgcgac ctcgttttag tctacgttta tctgtcttta
    14041 cttaatgtcc tttgttacag gccagaaagc ataactggcc tgaatattct ctctgggccc
    14101 actgttccac ttgtatcgtc ggtctgataa tcagactggg accacggtcc cactcgtatc
    14161 gtcgg
//
