LOCUS       40924_9701        7754 bp DNA     circular SYN 13-MAY-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7754)
  AUTHORS   Palmer AE, Giacomello M, Kortemme T, Hires SA, Lev-Ram V, Baker D, 
            Tsien RY
  TITLE     Ca2+ indicators based on computationally redesigned 
            calmodulin-peptide pairs.
  JOURNAL   Chem Biol. 2006 May;13(5):521-30.
  PUBMED    16720273
REFERENCE   2  (bases 1 to 7754)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 7754)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Chem
            Biol."; date: "2006-05"; volume: "13(5)"; pages: "521-30"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7754
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(86..943)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(944..1048)
                     /label=AmpR promoter
     primer_bind     complement(1123..1142)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable
                     marker"
     enhancer        1314..1693
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1694..1897
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        1942..1960
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1974..2048
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             2070..2144
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             2166..2240
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             2262..2336
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MSVLTPLLLRGLTGSARRLPVPRAK"
     CDS             2364..3047
                     /codon_start=1
                     /label=CFP
                     /note="cyan variant of GFP"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAA"
     CDS             3588..4322
                     /codon_start=1
                     /product="circularly permuted form of the Venus yellow
                     fluorescent protein (Nagai et al., 2004)"
                     /label=cp173Venus
                     /note="efficient FRET acceptor for CFP"
                     /translation="MDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRD
                     HMVLLEFVTAAGITLGMDELYKGGSGGMVSKGEELFTGVVPILVELDGDVNGHKFSVSG
                     EGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPE
                     GYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHN
                     VYITADKQKNGIKANFKIRHNIE"
     promoter        complement(4386..4404)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     polyA_signal    4430..4654
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4700..5128
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        5142..5471
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             5538..6329
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    6506..6639
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6676..6692)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(6700..6716)
                     /label=lac operator
                     /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(6724..6754)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(6769..6790)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     primer_bind     complement(6907..6924)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7078..7666)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
       61 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
      121 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
      181 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
      241 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
      301 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
      361 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
      421 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
      481 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
      541 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
      601 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
      661 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
      721 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
      781 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
      841 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
      901 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
      961 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     1021 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     1081 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     1141 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     1201 gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct
     1261 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg
     1321 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     1381 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     1441 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     1501 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     1561 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     1621 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     1681 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     1741 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     1801 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     1861 taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac
     1921 tgcttactgg cttatcgaaa ttaatacgac tcactatagg gagacccaag cttatgtccg
     1981 tcctgacgcc gctgctgctg cggggcttga caggctcggc ccggcggctc ccagtgccgc
     2041 gcgccaagat ccattcgttg ggggatccca tgtccgtcct gacgccgctg ctgctgcggg
     2101 gcttgacagg ctcggcccgg cggctcccag tgccgcgcgc caagatccat tcgttgggga
     2161 agcttatgtc cgtcctgacg ccgctgctgc tgcggggctt gacaggctcg gcccggcggc
     2221 tcccagtgcc gcgcgccaag atccattcgt tgggggatcc catgtccgtc ctgacgccgc
     2281 tgctgctgcg gggcttgaca ggctcggccc ggcggctccc agtgccgcgc gccaagatcc
     2341 attcgttggg gaagcttgcc accatggtga gcaagggcga ggagctgttc accggggtgg
     2401 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caggttcagc gtgtccggcg
     2461 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     2521 agctgcccgt gccctggccc accctcgtga ccaccctgac ctggggcgtg cagtgcttca
     2581 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     2641 acgtccagga gcgtaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     2701 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     2761 aggacggcaa catcctgggg cacaagctgg agtacaacta catcagccac aacgtctata
     2821 tcaccgccga caagcagaag aacggcatca aggcccactt caagatccgc cacaacatcg
     2881 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     2941 ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca
     3001 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgcccgc atgcatgacc
     3061 aactgacaga agagcagatt gcagagttca aagaagcctt ctcattattg gacaaggatg
     3121 gggacggcac catcaccaca aaggaacttg gcaccgctct gaggtcgctt ggacaaaacc
     3181 caacggaagc agaattgcag gatatgatca atgaagtcga tgctgatggc aatggaacga
     3241 tttactttcc tgaatttctt actatgatgg ctagaaaaat gaaggacaca gacagcgaag
     3301 aggaaatccg agaagcattc cgtgtttttg acaaggatgg gaacggctac atcagcgctg
     3361 ctgaattacg tcacgtcatg acaaacctcg gggagaagtt aacagatgaa gaagttgatg
     3421 aaatgataag ggaagcagat atcgatggtg atggccaagt aaactatgaa gagtttgtac
     3481 aaatgatgac agcaaagggg gggaagaggc gctggcagaa aacaggccat gctgtccgag
     3541 cattcggaag actgaagaag atctccagct ccggggcact ggagctcatg gacggcggcg
     3601 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     3661 ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac gagaagcgcg
     3721 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     3781 tgtacaaggg tggcagcggt ggcatggtga gcaagggcga ggagctgttc accggggtgg
     3841 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     3901 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gctgatctgc accaccggca
     3961 agctgcccgt gccctggccc accctcgtga ccaccctggg ctacggcctg cagtgcttcg
     4021 cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     4081 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     4141 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     4201 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     4261 tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg
     4321 agtaagaatt ctgcagatat ccatcacact ggcggccgct cgagcatgca tctagagggc
     4381 cctattctat agtgtcacct aaatgctaga gctcgctgat cagcctcgac tgtgccttct
     4441 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     4501 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     4561 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     4621 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     4681 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4741 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4801 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4861 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     4921 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4981 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     5041 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5101 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     5161 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     5221 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     5281 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     5341 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5401 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     5461 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag
     5521 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     5581 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     5641 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     5701 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     5761 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     5821 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     5881 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     5941 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     6001 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     6061 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     6121 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     6181 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     6241 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     6301 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     6361 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     6421 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     6481 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     6541 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     6601 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     6661 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     6721 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     6781 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     6841 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     6901 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     6961 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     7021 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     7081 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     7141 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     7201 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     7261 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     7321 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     7381 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     7441 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     7501 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     7561 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     7621 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     7681 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     7741 gagattatca aaaa
//
