LOCUS       sequence-200539-        3932 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Plasmid encoding 2ndVal mKate2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3932)
  AUTHORS   Balleza E, Kim JM, Cluzel P
  TITLE     Systematic characterization of maturation time of fluorescent 
            proteins in living cells.
  JOURNAL   Nat Methods. 2018 Jan;15(1):47-51. doi: 10.1038/nmeth.4509. Epub 
            2017 Nov 20.
  PUBMED    29320486
REFERENCE   2  (bases 1 to 3932)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/nmeth.4509"; journalName: "Nat Methods"; date: "2018-01"; 
            volume: "15"; issue: "1"; pages: "47-51"
COMMENT     Sequence Label: pEB2-2ndVal mKate2
FEATURES             Location/Qualifiers
     source          1..3932
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     77..95
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             280..978
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     terminator      1016..1062
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1235..2185)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2233..2455
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(2961..2995)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     CDS             complement(3036..3830)
                     /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(3147..3166)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3757..3776
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 aaccttacca gagggcgccc cagctggcaa ttccgacgtc taagaaacca ttattatcat
       61 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcacc acagctaaca
      121 ccacgtcgtc cctatctgct gccctaggtc tatgagtggt tgctggataa ctttacgggc
      181 atgcataagg ctcgtatgat atattcaggg agaccacaac ggtttccctc tacaaataat
      241 tttgtttaac ttttctagat ttaagaagga gatatacata tggtgtctga gctgattaag
      301 gagaacatgc acatgaagct gtacatggag ggcaccgtga acaaccacca cttcaagtgc
      361 acatccgagg gcgaaggcaa gccctacgag ggcacccaga ccatgagaat caaggcggtc
      421 gagggcggcc ctttaccctt cgccttcgac atcctggcta ccagcttcat gtacggcagc
      481 aaaaccttca tcaaccacac ccagggcatc cccgacttct ttaagcagtc cttccctgag
      541 ggcttcacat gggagagagt caccacatac gaagacgggg gcgtgctgac cgctacccag
      601 gacaccagct tacaggacgg ctgcttgatc tacaacgtca agatcagagg ggtgaacttc
      661 ccatccaacg gccctgtgat gcagaagaaa acattaggct gggaggcctc caccgagacc
      721 ctgtaccccg ctgacggcgg cctggaaggc agagccgaca tggccctgaa gttagtgggc
      781 gggggccacc tgatctgcaa cttgaagacc acatacagat ccaagaaacc cgctaagaac
      841 ttgaagatgc ccggcgtcta ctatgtggac agaagactgg aaagaatcaa ggaggccgac
      901 aaagagacct acgtcgagca gcacgaggtg gctgtggcca gatactgcga cctgcctagc
      961 aaactggggc acagataaat gtccagacct gcaggcatgc aagctctaga ggcatcaaat
     1021 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgaa atgcaccaaa
     1081 aactcgtaaa agctctgatg tatctatctt ttttacaccg ttttcatctg tgcatatgga
     1141 cagttttccc tttgatatct aacggtgaac agttgttcta cttttgtttg ttagtcttga
     1201 tgcttcactg atagatacaa gagccataag aacctcagat ccttccgtat ttagccagta
     1261 tgttctctag tgtggttcgt tgtttttgcg tgagccatga gaacgaacca ttgagatcat
     1321 gcttactttg catgtcactc aaaaattttg cctcaaaact ggtgagctga atttttgcag
     1381 ttaaagcatc gtgtagtgtt tttcttagtc cgttacgtag gtaggaatct gatgtaatgg
     1441 ttgttggtat tttgtcacca ttcattttta tctggttgtt ctcaagttcg gttacgagat
     1501 ccatttgtct atctagttca acttggaaaa tcaacgtatc agtcgggcgg cctcgcttat
     1561 caaccaccaa tttcatattg ctgtaagtgt ttaaatcttt acttattggt ttcaaaaccc
     1621 attggttaag ccttttaaac tcatggtagt tattttcaag cattaacatg aacttaaatt
     1681 catcaaggct aatctctata tttgccttgt gagttttctt ttgtgttagt tcttttaata
     1741 accactcata aatcctcata gagtatttgt tttcaaaaga cttaacatgt tccagattat
     1801 attttatgaa tttttttaac tggaaaagat aaggcaatat ctcttcacta aaaactaatt
     1861 ctaatttttc gcttgagaac ttggcatagt ttgtccactg gaaaatctca aagcctttaa
     1921 ccaaaggatt cctgatttcc acagttctcg tcatcagctc tctggttgct ttagctaata
     1981 caccataagc attttcccta ctgatgttca tcatctgagc gtattggtta taagtgaacg
     2041 ataccgtccg ttctttcctt gtagggtttt caatcgtggg gttgagtagt gccacacagc
     2101 ataaaattag cttggtttca tgctccgtta agtcatagcg actaatcgct agttcatttg
     2161 ctttgaaaac aactaattca gacatacatc tcaattggtc taggtgattt taatcactat
     2221 accaattgag atgggctagt caatgataat tactagtcct tttcctttga gttgtgggta
     2281 tctgtaaatt ctgctagacc tttgctggaa aacttgtaaa ttctgctaga ccctctgtaa
     2341 attccgctag acctttgtgt gttttttttg tttatattca agtggttata atttatagaa
     2401 taaagaaaga ataaaaaaag ataaaaagaa tagatcccag ccctgtgtat aactcactac
     2461 tttagtcagt tccgcagtat tacaaaagga tgtcgcaaac gctgtttgct cctctacaaa
     2521 acagacctta aaaccctaaa ggcttaagta gcaccctcgc aagctcgggc aaatcgctga
     2581 atattccttt tgtctccgac catcaggcac ctgagtcgct gtctttttcg tgacattcag
     2641 ttcgctgcgc tcacggctct ggcagtgaat gggggtaaat ggcactacag gcgcctttta
     2701 tggattcatg caaggaaact acccataata caagaaaagc ccgtcacggg cttctcaggg
     2761 cgttttatgg cgggtctgct atgtggtgct atctgacttt ttgctgttca gcagttcctg
     2821 ccctctgatt ttccagtctg accacttcgg attatcccgt gacaggtcat tcagactggc
     2881 taatgcaccc agtaaggcag cggtatcatc aacaggctta cccgtcttac tgtcaaccag
     2941 acccgccagg attctcatgc ataaaaaacg cccggcggca accgagcgtt ctgaattaat
     3001 taatcatcgc gactgcagag atcccagagt ccttattaga agaactcgtc aagaaggcga
     3061 tagaaggcga tgcgctgcga atcgggagcg gcgataccgt aaagcacgag gaagcggtca
     3121 gcccattcgc cgccaagctc ttcagcaata tcacgggtag ccaacgctat gtcctgatag
     3181 cggtccgcca cacccagccg gccacagtcg atgaatccag aaaagcggcc attttccacc
     3241 atgatattcg gcaagcaggc atcgccatgg gtcacgacga gatcctcgcc gtcgggcatg
     3301 cgcgccttga gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc ttcgtccaga
     3361 tcatcctgat cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat gcgatgtttc
     3421 gcttggtggt cgaatgggca ggtagccgga tcaagcgtat gcagccgccg cattgcatca
     3481 gccatgatgg atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc
     3541 acttcgccca atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagctgcg
     3601 caaggaacgc ccgtcgtggc cagccacgat agccgcgctg cctcgtcctg cagttcattc
     3661 agggcaccgg acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc tgacagccgg
     3721 aacacggcgg catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc
     3781 tccacccaag cggccggaga acctgcgtgc aatccatctt gttcaatcat gcgaaacgat
     3841 cctcatcctg tctcttgatc agatcttgat cccctgcgcc atcagatcct tggcggcaag
     3901 aaagccatcc agtttacttt gcagggcttc cc
//
