LOCUS       sequence-459823-        2963 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MicF RNA toehold switch with sfgfp reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2963)
  AUTHORS   Diaz KJ, Jarquin J, Petrosyan A, Takahashi MK
  TITLE     Improved RNA toehold switch sensitivity using the NanoLuc 
            complementation reporter.
  JOURNAL   MicroPubl Biol. 2025 Jan 2;2025:10.17912/micropub.biology.001334. 
            doi: 10.17912/micropub.biology.001334. eCollection 2025.
  PUBMED    39839716
REFERENCE   2  (bases 1 to 2963)
  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: "MicroPubl 
            Biol."; date: "2025-01-2"; volume: 
            "2025:10.17912/micropub.biology.001334. doi"; pages: " 
            10.17912/micropub.biology.001334. eCollection 2025"
COMMENT     Sequence Label: MKT071
FEATURES             Location/Qualifiers
     source          1..2963
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     179..197
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        224..258
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     CDS             357..1070
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             357..1067
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(384..412)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1007..1027
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1117..1203
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(1368..1956)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1448..1467)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2044..2138)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2169..2963)
                     /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(2280..2299)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2890..2909
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 gcgaaacgat cctcatcctg tctcttgatc agatcttgat cccctgcgcc atcagatcct
       61 tggcggcaag aaagccatcc agtttacttt gcagggcttc ccaaccttac cagagggcgc
      121 cccagctggc aattccgacg tctaagaaac cattattatc atgacattaa cctataaaaa
      181 taggcgtatc acgaggccct ttcgtcttca cctcgaggta cccttgacag ctagctcagt
      241 cctaggtata atactagtgt gaatgacggt aataaataaa gttaatgatg atagcggact
      301 ttagaacaga ggagataaag atggctatca tcatgaacct ggcggcagcg caaaagagca
      361 aaggagaaga acttttcact ggagttgtcc caattcttgt tgaattagat ggtgatgtta
      421 atgggcacaa attttctgtc cgtggagagg gtgaaggtga tgctacaaac ggaaaactca
      481 cccttaaatt tatttgcact actggaaaac tacctgttcc gtggccaaca cttgtcacta
      541 ctctgaccta tggtgttcaa tgcttttccc gttatccgga tcacatgaaa cggcatgact
      601 ttttcaagag tgccatgccc gaaggttatg tacaggaacg cactatatct ttcaaagatg
      661 acgggaccta caagacgcgt gctgaagtca agtttgaagg tgataccctt gttaatcgta
      721 tcgagttaaa gggtattgat tttaaagaag atggaaacat tcttggacac aaactcgagt
      781 acaactttaa ctcacacaat gtatacatca cggcagacaa acaaaagaat ggaatcaaag
      841 ctaacttcaa aattcgccac aacgttgaag atggttccgt tcaactagca gaccattatc
      901 aacaaaatac tccaattggc gatggccctg tccttttacc agacaaccat tacctgtcga
      961 cacaatctgt cctttcgaaa gatcccaacg aaaagcgtga ccacatggtc cttcttgagt
     1021 ttgtaactgc tgctgggatt acacatggca tggatgagct ctacaaataa taaggtgctt
     1081 aataaggatc ctgtcgatcg cgcgtgctag aggcatcaaa taaaacgaaa ggctcagtcg
     1141 aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcct gagtaggaca
     1201 aatccgccgc cctagaccta gggcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     1261 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     1321 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     1381 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     1441 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     1501 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     1561 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     1621 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1681 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     1741 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     1801 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     1861 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     1921 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     1981 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gactagtgct
     2041 tggattctca ccaataaaaa acgcccggcg gcaaccgagc gttctgaaca aatccagatg
     2101 gagttctgag gtcattactg gatctatcaa caggagtcca agcgagctct cgaaccccag
     2161 agtcccgctc agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga
     2221 gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca
     2281 atatcacggg tagccaacgc tatgtcctga tagcggtccg ccacacccag ccggccacag
     2341 tcgatgaatc cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca
     2401 tgggtcacga cgagatcctc gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg
     2461 gctggcgcga gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc
     2521 atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc
     2581 ggatcaagcg tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga
     2641 gcaaggtgag atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt
     2701 cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac
     2761 gatagccgcg ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca
     2821 aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt
     2881 gtctgttgtg cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg
     2941 tgcaatccat cttgttcaat cat
//
