LOCUS       pET_Syt1_C2AB_14        6923 bp DNA     circular SYN 15-JUN-2023
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6923)
  AUTHORS   nmehta22
  TITLE     Direct Submission
  JOURNAL   Exported Sep 12, 2023 from SnapGene 7.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6923
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     CDS             complement(548..1363)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /note="color: #ccffcc"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      1485..2073
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(2500..2691)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /note="color: #993366"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3500..4582)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /note="color: #993366"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(4583..4660)
                     /gene="lacI"
                     /label=lacI promoter
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     4874..4922
                     /label=BL14
                     /note="This oligo (used with BL13) will replace the linker 
                     MDELYK to GSSGSS in monomeric superfolder GFP, CFP and 
                     YFP."
                     /note="color: black; sequence: 
                     GATCTAGAGTCGCGGCCGCTTTAcccgctggagccggagctGCCGTGAGTGATCCCGGC
                     "
     promoter        4969..4987
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    4988..5012
                     /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)."
                     /note="color: #31849b"
     RBS             5042..5048
                     /note="ribosome binding site"
                     /note="color: #a6acb3"
     CDS             5057..5059
                     /codon_start=1
                     /product="start codon"
                     /label=ATG
                     /note="color: #993366"
                     /translation="M"
     CDS             5069..5086
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHH"
     CDS             5096..5113
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /note="color: #cc99b2
                     Cleavage site after base 5108"
                     /translation="LVPRGS"
     CDS             5120..5956
                     /codon_start=1
                     /label=rSyt1_FL
                     /note="color: #993366"
                     /translation="GKLQYSLDYDFQNNQLLVGIIQAAELPALDMGGTSDPYVKVFLLP
                     DKKKKFETKVHRKTLNPVFNEQFTFKVPYSELGGKTLVMAVYDFDRFSKHDIIGEFKVP
                     MNTVDFGHVTEEWRDLQSAEKEEQEKLGDICFSLRYVPTAGKLTVVILEAKNLKKMDVG
                     GLSDPYVKIHLMQNGKRLKKKKTTIKKNTLNPYYNESFSFEVPFEQIQKVQVVVTVLDY
                     DKIGKNDAIGKVFVGYNSTGAELRHWSDMLANPRRPIAQWHTLQVEEEVDAMLAVKK"
     misc_feature    5120..5956
                     /label=Syt1 C2AB 80-421
                     /note="color: #a6acb3"
     primer_bind     5347..5370
                     /label=Primer 1
                     /note="color: black; sequence: attaggtggcaaaaccctggtgat; 
                     added: 2020-09-16"
     primer_bind     complement(5347..5370)
                     /label=Primer 2
                     /note="color: black; sequence: atcaccagggttttgccacctaat; 
                     added: 2020-09-16"
     primer_bind     5936..6021
                     /label=msGFP-FLAG 60 F (EC13459)
                     /note="color: black; sequence: 
                     GCCATGCTGGCTGTCAAGAAGGGATCCGGAAGTTCGGGGTCCAGCGGTTCTTCAGGCAG
                     TTCCATGGATAGCACTGAGAGCCTGTT; added: 2020-09-21"
     primer_bind     complement(5937..5956)
                     /label=Syt1 C2AB 80 R
                     /note="color: black; sequence: 
                     ATAAATACCTAAGCTTttacttcttgacagccagcatgg; added: 2020-09-04"
     CDS             5957..5998
                     /label=GS Linker
                     /note="color: #993366; direction: RIGHT"
     misc_feature    5957..5962
                     /label=linker for pre-prolactin
                     /note="found in original sequence"
                     /note="color: #a6acb3"
     primer_bind     5997..6034
                     /label=DSTES
                     /note="Use together with BL14 to amplify msGFP, msYFP, and 
                     msCFP with the improved N-terminal codons from A24. 
                     Subclone the PCR product back into the same vector using 
                     BamHI and NotI."
                     /note="color: black; sequence: 
                     CCGGGATCCACCGGTCGCCACCATGGATAGCACTGAGAGCCTGTTCACCGGGGTGGTG"
     CDS             5999..6694
                     /codon_start=1
                     /label=msGFP
                     /note="color: #05fd14"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHG"
     CDS             5999..6016
                     /codon_start=1
                     /product="""An N-terminal MDSTES peptide in place of 
                     MVSKGEE """"In particular, the widely used red FP mCherry 
                     contains the N- and C-terminal peptides of EGFP (12), and 
                     we find that the N-terminal peptide of EGFP can promote 
                     cytotoxicity of mCherry during bacterial expression.""
                       A photostable monomeric superfolder GFPFernando Valbuena,
                     Ivy Fizgerald, Rita L. Strack, Neal Andruska, Luke Smith, 
                     Benjamin S. Glick 
                     "
                     /label=N-terminus mutation
                     /note="color: #993366"
                     /translation="MDSTES"
     primer_bind     6002..6025
                     /label=new N GFP lin F
                     /note="color: black; sequence: 
                     GCCATTTGCTAAACCTGTTAGAGCTGATAGCACTGAGAGCCTGTTCACC"
     CDS             6086..6088
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850."
                     /label=superfolder mutation (1/8) S30R
                     /note="color: #993366"
                     /translation="R"
     CDS             6113..6115
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850."
                     /label=superfolder mutation (2/8) Y39N
                     /note="color: #993366"
                     /translation="N"
     primer_bind     complement(6126..6148)
                     /label=msGFP N term R
                     /note="color: black; sequence: GGTGGTGCAGATGAACTTCAGGG; 
                     added: 2020-08-31"
     CDS             6293..6295
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850."
                     /label=superfolder mutation (3/8) F99T
                     /note="color: #993366"
                     /translation="T"
     CDS             6311..6313
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850."
                     /label=superfolder mutation (4/8) N105T
                     /note="color: #993366"
                     /translation="T"
     CDS             6431..6433
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850. 
                     Reversion of this mutation (F145Y) restores photostability 
                     of superfolding mutant with only a minor effect on folding 
                     Pedelacq JD, Cabantous S, Tran T, Terwilliger TC, Waldo GS.
                     Engineering and characterization of a superfolder green 
                     fluorescent protein. Nat Biotechnol 2006;24:79–88.
                     "
                     /label=superfolder mutation (5/8) Y145F
                     /note="color: #993366"
                     /translation="F"
     CDS             6455..6457
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850."
                     /label=superfolder mutation (6/8) M153T
                     /note="color: #993366"
                     /translation="T"
     primer_bind     complement(6478..6502)
                     /label=BQ13
                     /note="color: black; sequence: GCGGATCTTGAAGTTGGCCTTGATG"
     CDS             6485..6487
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850"
                     /label=superfolder mutation (7/8) V163A
                     /note="color: #993366"
                     /translation="A"
     CDS             6509..6511
                     /codon_start=1
                     /product="Shagin DA, Barsova EV, Yanushevich YG, Fradkov 
                     AF, Lukyanov KA, Labas YA, Semenova TN, Ugalde JA, Meyers 
                     A, Nunez JM, Widder EA, Lukyanov SA, Matz MV. GFP-like 
                     proteins as ubiquitous metazoan superfamily: evolution of 
                     functional features and structural complexity. Mol Biol 
                     Evol 2004;21(5):841–850."
                     /label=superfolder mutation (8/8) I171V
                     /note="color: #993366"
                     /translation="V"
     primer_bind     6569..6584
                     /label=msGFP C term F
                     /note="color: black; sequence: GGCCCCGTGCTGCTGC; added: 
                     2020-08-31"
     CDS             6614..6616
                     /codon_start=1
                     /product="Costantini LM, Fossati M, Francolini M, Snapp EL.
                     Assessing the tendency of fluorescent proteins to 
                     oligomerize under physiologic conditions. Traffic 
                     2012;13(5):643–649.
                     Zacharias DA, Violin JD, Newton AC, Tsien RY. Partitioning 
                     of lipid-modified monomeric GFPs into membrane microdomains
                     of live cells. Science 2002;296:913–916.
                     "
                     /label=monomerizing mutation A206K
                     /note="color: #993366"
                     /translation="K"
     primer_bind     complement(6677..6694)
                     /label=BL14
                     /note="This oligo (used with BL13) will replace the linker 
                     MDELYK to GSSGSS in monomeric superfolder GFP, CFP and 
                     YFP."
                     /note="color: black; sequence: 
                     GATCTAGAGTCGCGGCCGCTTTAcccgctggagccggagctGCCGTGAGTGATCCCGGC
                     "
     primer_bind     complement(6677..6694)
                     /label=msGFP FLAG 60 R (EC13899)
                     /note="color: black; sequence: 
                     GATAAGCTTGATATCGAATTTTACTTGTCGTCATCGTCTTTGTAGTCTGAAGAACCGCC
                     GTGAGTGATCCCGGC; added: 2020-09-21"
     primer_bind     complement(6684..6694)
                     /label=BL24
                     /note="This oligo is used to amplify msfFPs (Green, Cyan, 
                     Yellow) from the pMSGFP vector into the NT-HT vector for 
                     the Tub1 fussion.  The cut sites are BspEI for the 
                     N-terminal and SacI for the C-terminal. Use this oligo with
                     BL23."
                     /note="color: black; sequence: 
                     GGTTCCAGAGGGAGCACCGGAGCTCGTcccgctggagccggagctGCCGTGAGTGA"
     CDS             6689..6691
                     /codon_start=1
                     /product="Tsien RY. The green fluorescent protein. Annu Rev
                     Biochem 1998;67:509–544."
                     /label=monomeric mutation L231H
                     /note="color: #993366"
                     /translation="H"
     primer_bind     complement(6695..6713)
                     /label=Syt1 C2AB 80 R
                     /note="color: black; sequence: 
                     ATAAATACCTAAGCTTttacttcttgacagccagcatgg; added: 2020-09-04"
     CDS             6756..6773
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHH"
     primer_bind     complement(6823..6842)
                     /label=T7 Reverse
                     /note="color: black; sequence: TAGTTATTGCTCAGCGGTGG"
     terminator      6840..6887
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
ORIGIN
        1 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
       61 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
      121 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
      181 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
      241 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgaa
      541 ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat
      601 tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc
      661 agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa
      721 tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag
      781 tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa
      841 caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc
      901 gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag
      961 gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat
     1021 caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc
     1081 atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca
     1141 gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt
     1201 tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt
     1261 gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta
     1321 atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac
     1381 tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg tgagttttcg
     1441 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     1501 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     1561 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     1621 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     1681 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     1741 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     1801 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     1861 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     1921 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     1981 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     2041 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     2101 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     2161 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     2221 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt
     2281 acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc tgctctgatg
     2341 ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca tggctgcgcc
     2401 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     2461 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     2521 accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa gcgattcaca
     2581 gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg ttaatgtctg
     2641 gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca ctgatgcctc
     2701 cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag agaggatgct
     2761 cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg agggtaaaca
     2821 actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat gccagcgctt
     2881 cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga tgcagatccg
     2941 gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa cacggaaacc
     3001 gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt cgcttcacgt
     3061 tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc agcctagccg
     3121 ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc cggcgataat
     3181 ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt gagcgagggc
     3241 gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg
     3301 gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt gcatgataaa
     3361 gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga aggagctgac
     3421 tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag tgagctaact
     3481 tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     3541 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg
     3601 tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag
     3661 agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg
     3721 tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga
     3781 tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct
     3841 gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt
     3901 gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat
     3961 tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg
     4021 ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc
     4081 gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa
     4141 gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca
     4201 gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt
     4261 tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat
     4321 cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg
     4381 tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa
     4441 tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc
     4501 tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat
     4561 cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc
     4621 atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc
     4681 ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc
     4741 gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc cacggggcct
     4801 gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc ccgatcttcc
     4861 ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg
     4921 gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaatta atacgactca
     4981 ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg tttaacttta
     5041 agaaggagat atacatatgg gcagcagcca tcatcatcat catcacggca gcggcctggt
     5101 gccgcgcggc agcgctagcg gaaagctcca atattcactg gactatgact tccagaataa
     5161 ccagctgttg gtgggaatca tccaggctgc tgaactgccc gccctggaca tggggggtac
     5221 atccgatcca tacgtcaaag tcttcctgct gcctgacaaa aagaagaaat ttgagactaa
     5281 agtccaccgg aaaaccctca atccagtctt caatgaacaa tttactttca aggtacccta
     5341 ctcggaatta ggtggcaaaa ccctggtgat ggctgtgtat gactttgatc gcttctccaa
     5401 gcacgacatc atcggagagt tcaaagttcc tatgaacacc gtggattttg gccatgtgac
     5461 cgaggagtgg cgcgatctcc agagcgctga gaaagaagag caagagaaac tgggtgacat
     5521 ctgcttctcc ctccgctacg tccctactgc cggcaaactg actgttgtca ttctggaagc
     5581 caagaacctg aagaagatgg atgtgggtgg cttatctgat ccctacgtga agattcacct
     5641 gatgcagaac ggtaagaggc tgaagaagaa aaagacgacg attaagaaga acacactcaa
     5701 cccctactac aacgagtcct tcagctttga agttccgttc gagcaaatcc agaaagtgca
     5761 agtggtggta actgttttgg actatgacaa gattggcaag aacgacgcca tcggaaaagt
     5821 cttcgttggt tacaacagca ctggggcgga gctgcgacac tggtcagaca tgctggccaa
     5881 cccccggcga cccatcgcac agtggcacac tctgcaggta gaggaggagg ttgatgccat
     5941 gctggctgtc aagaagggat ccggaagttc ggggtccagc ggttcttcag gcagttccat
     6001 ggatagcact gagagcctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     6061 cgtaaacggc cacaagttca gcgtgcgcgg cgagggcgag ggcgatgcca ccaacggcaa
     6121 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     6181 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     6241 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcaccttcaa
     6301 ggacgacggc acctacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     6361 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     6421 ggagtacaac ttcaacagcc acaacgtcta tatcaccgcc gacaagcaga agaacggcat
     6481 caaggccaac ttcaagatcc gccacaacgt ggaggacggc agcgtgcagc tcgccgacca
     6541 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     6601 gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     6661 ggagttcgtg accgccgccg ggatcactca cggctaaaag cttaggtatt tattcggcgc
     6721 aaagtgcgtc gggtgatgct gccaacttag tcgagcacca ccaccaccac cactgagatc
     6781 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
     6841 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
     6901 ctatatccgg attggcgaat ggg
//
