LOCUS       sequence-199485-        2483 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro expression of mKalama1 from the T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2483)
  AUTHORS   Stark JC, Huang A, Nguyen PQ, Dubner RS, Hsu KJ, Ferrante TC, 
            Anderson M, Kanapskyte A, Mucha Q, Packett JS, Patel P, Patel R, Qaq
            D, Zondor T, Burke J, Martinez T, Miller-Berry A, Puppala A, 
            Reichert K, Schmid M, Brand L, Hill LR, Chellaswamy JF, Faheem N, 
            Fetherling S, Gong E, Gonzalzles EM, Granito T, Koritsaris J, Nguyen
            B, Ottman S, Palffy C, Patel A, Skweres S, Slaton A, Woods T, 
            Donghia N, Pardee K, Collins JJ, Jewett MC
  TITLE     BioBits Bright: A fluorescent synthetic biology education kit.
  JOURNAL   Sci Adv. 2018 Aug 1;4(8):eaat5107. doi: 10.1126/sciadv.aat5107. 
            eCollection 2018 Aug.
  PUBMED    30083609
REFERENCE   2  (bases 1 to 2483)
  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: "Sci Adv."; 
            date: "2018-08-1"; volume: "4(8):eaat5107. doi"; pages: " 
            10.1126/sciadv.aat5107. eCollection 2018 Aug"
COMMENT     Sequence Label: pJL1-mKalama1
FEATURES             Location/Qualifiers
     source          1..2483
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             706..728
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(759..778)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(760..778)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(806..1394)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(886..905)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(1505..1524)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1516..2331
                     /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="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1589..1608)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      complement(2349..2396)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     2392..2410
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(join(2462..2483,1..692,693..695,696..698))
                     /codon_start=1
                     /product="photostable monomeric blue variant of GFP (Ai et 
                     al., 2007)"
                     /label=mKalama1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVEMDGDVNGRKFSVRGVGEGDATHGKLTL
                     KFICTSGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GSYKTRAEVKFEGDTLVNRIVLKGTDFKEDGNILGHKLEYNMNVGNVYITADKQKNGIK
                     ANFEIRHNVEDGGVQLADHYQQNTPIGDGSVLLPDNHYLSVQVKLSKDPNEKRDHMVLL
                     EFRTAAGITPGMDELYK"
ORIGIN
        1 ccgggagtga tcccggcggc ggtgcggaac tccagcagga ccatgtgatc gcgcttctcg
       61 ttggggtctt tgctcagctt cacctgcacg ctcaggtagt ggttgtcggg cagcagcacg
      121 gagccgtcgc cgatgggggt gttctgctgg tagtggtcgg cgagctgcac gccgccgtcc
      181 tcgacgttgt gcctgatctc gaagttcgct ttgatgccgt tcttctgctt gtcggccgtg
      241 atatagacgt tcccgacgtt catgttgtac tccagcttgt gccccaggat gttaccgtcc
      301 tccttgaagt cggtgccctt cagcacgatg cggttcacca gggtgtcgcc ctcgaacttc
      361 acctcggcgc gggtcttgta gctgccgtcg tccttgaaga agatggtgcg ctcctggacg
      421 tagccttcgg gcatggcgga cttgaagaag tcgtgctgct tcatgtggtc ggggtagcgg
      481 gagaagcact gcacgccgta agacagggtg gtcacgaggg taggccaggg cacgggcagc
      541 ttgccggagg tgcagatgaa cttcagggtc agcttgccgt gggtggcatc accctcgccc
      601 acgccgcgca cgctgaactt gcggccgttt acgtcgccgt ccatctcgac caggatgggc
      661 accaccccgg tgaacagctc ctcgcccttg ctcaccatat gtatatctcc ttcttaaagt
      721 taaacaaaat tatttctaga gggaaaccgt tgtggtctcc ctatagtgag tcgtattaat
      781 ttcgcgggat cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
      841 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
      901 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
      961 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     1021 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     1081 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     1141 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     1201 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     1261 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     1321 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     1381 aaaaaggatc tcaagaagat cctttgatct cacgttgtgt ctcaaaatct ctgatgttac
     1441 attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt
     1501 aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cttgctcgaa gccgcgatta
     1561 aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa
     1621 tcaggtgcga caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa
     1681 catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg
     1741 acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg
     1801 ttactcacca ctgcgatccc cgggaaaaca gcattccagg tattagaaga atatcctgat
     1861 tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct
     1921 gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga
     1981 atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt
     2041 gaacaagtct ggaaagaaat gcataagctt ttgccattct caccggattc agtcgtcact
     2101 catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt
     2161 gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc
     2221 ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat
     2281 cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta atcagaattg
     2341 gctttcagca aaaaacccct caagacccgt ttagaggccc caaggggtta tgctagttat
     2401 tgctcagcgg tggcagcagc caactcagct tcctttcggg ctttgttagc agccggtcga
     2461 cttacttgta cagctcgtcc atg
//
