LOCUS       sequence-68379-e        4041 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BIOFAB reporter plasmid for measuring T7 early termination 
            efficiency.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4041)
  AUTHORS   Mutalik VK, Guimaraes JC, Cambray G, Lam C, Christoffersen MJ, Mai 
            QA, Tran AB, Paull M, Keasling JD, Arkin AP, Endy D
  TITLE     Precise and reliable gene expression via standard transcription and 
            translation initiation elements.
  JOURNAL   Nat Methods. 2013 Apr;10(4):354-60. doi: 10.1038/nmeth.2404. Epub 
            2013 Mar 10.
  PUBMED    23474465
REFERENCE   2  (bases 1 to 4041)
  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"; doi: 
            "10.1038/nmeth.2404"; journalName: "Nat Methods"; date: "2013-04"; 
            volume: "10"; issue: "4"; pages: "354-60"
COMMENT     Sequence Label: pFAB803
FEATURES             Location/Qualifiers
     source          1..4041
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(320..338,339..344,345..361,362..367,368..393)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    320..338
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    345..363
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             407..418
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             425..1102
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      1206..1233
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             1350..1361
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(1368..1370,1371..2084)
                     /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="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             1368..2081
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1398..1426)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2021..2041
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      2117..2188
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2204..2231
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(2302..2319)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2436..2981)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(2516..2535)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(3095..3189)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3220..4014)
                     /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(3331..3350)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3941..3960
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 atcagatcat gatcccctgc gccatcagat ccttggcggc aagaaagcca tccagtttac
       61 tttgcagggc ttcccaacct taccagaggg cgccccagct ggcaattccg acgtctaggg
      121 ataacagggt aattacggcc ccagaattca aaagatctta agtaagtaag agtatacgta
      181 tatcggctaa aacgtattaa ggcgcttcgg cgcctttttt tatgggggta ttttcatccc
      241 aatccacacg tccaacgcac agcaaacacc acgtcgaccc tatcagctgc gtgctttcta
      301 tgagtcgttg ctgcatatct ccctatcagt gatagagatt gacatcccta tcagtgatag
      361 agatactgag cacatcagca ggacgcactg acctttggaa ttcattaaag aggagaaagg
      421 taccatggcg agtagcgaag acgttatcaa agagttcatg cgtttcaaag ttcgtatgga
      481 aggttccgtt aacggtcacg agttcgaaat cgaaggtgaa ggtgaaggtc gtccgtacga
      541 aggtacccag accgctaaac tgaaagttac caaaggtggt ccgctgccgt tcgcttggga
      601 catcctgtcc ccgcagttcc agtacggttc caaagcttac gttaaacacc cggctgacat
      661 cccggactac ctgaaactgt ccttcccgga aggtttcaaa tgggaacgtg ttatgaactt
      721 cgaagacggt ggtgttgtta ccgttaccca ggactcctcc ctgcaagacg gtgagttcat
      781 ctacaaagtt aaactgcgtg gtaccaactt cccgtccgac ggtccggtta tgcagaaaaa
      841 aaccatgggt tgggaagctt ccaccgaacg tatgtacccg gaagacggtg ctctgaaagg
      901 tgaaatcaaa atgcgtctga aactgaaaga cggtggtcac tacgacgctg aagttaaaac
      961 cacctacatg gctaaaaaac cggttcagct gccgggtgct tacaaaaccg acatcaaact
     1021 ggacatcacc tcccacaacg aagactacac catcgttgaa cagtacgaac gtgctgaagg
     1081 tcgtcactcc accggtgctt aataatctgc tgagtgatag actcaaggtc gctcctagcg
     1141 agtggccttt atgattatca ctttacttat gtgatacaac cctcaagaga aaatgtaatc
     1201 acactggctc accttcgggt gggcctttct gcgtttataa ggagacactt tatgtttaag
     1261 aagggcggac tcactataga gggacaaact caaggtcatt cgcaagagtg gcctttatga
     1321 ttgaccttct tccggttaat caattcatta aagaggagaa aggtaccatg agcaaaggag
     1381 aagaactttt cactggagtt gtcccaattc ttgttgaatt agatggtgat gttaatgggc
     1441 acaaattttc tgtccgtgga gagggtgaag gtgatgctac aaacggaaaa ctcaccctta
     1501 aatttatttg cactactgga aaactacctg ttccgtggcc aacacttgtc actactctga
     1561 cctatggtgt tcaatgcttt tcccgttatc cggatcacat gaaacggcat gactttttca
     1621 agagtgccat gcccgaaggt tatgtacagg aacgcactat atctttcaaa gatgacggga
     1681 cctacaagac gcgtgctgaa gtcaagtttg aaggtgatac ccttgttaat cgtatcgagt
     1741 taaagggtat tgattttaaa gaagatggaa acattcttgg acacaaactc gagtacaact
     1801 ttaactcaca caatgtatac atcacggcag acaaacaaaa gaatggaatc aaagctaact
     1861 tcaaaattcg ccacaacgtt gaagatggtt ccgttcaact agcagaccat tatcaacaaa
     1921 atactccaat tggcgatggc cctgtccttt taccagacaa ccattacctg tcgacacaat
     1981 ctgtcctttc gaaagatccc aacgaaaagc gtgaccacat ggtccttctt gagtttgtaa
     2041 ctgctgctgg gattacacat ggcatggatg agctctacaa ataaggatcg gttgtcgagt
     2101 aaggatctcc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg cctttcgttt
     2161 tatctgttgt ttgtcggtga acgctctcta ctagagtcac actggctcac cttcgggtgg
     2221 gcctttctgc gtttatagga tcctaactcg agctgaagcc aatagggata acagggtaat
     2281 cctagggata tattccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg
     2341 gcgagcggaa atggcttacg aacggggcgg agatttcctg gaagatgcca ggaagatact
     2401 taacagggaa gtgagagggc cgcggcaaag ccgtttttcc ataggctccg cccccctgac
     2461 aagcatcacg aaatctgacg ctcaaatcag tggtggcgaa acccgacagg actataaaga
     2521 taccaggcgt ttccccctgg cggctccctc gtgcgctctc ctgttcctgc ctttcggttt
     2581 accggtgtca ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga cactcagttc
     2641 cgggtaggca gttcgctcca agctggactg tatgcacgaa ccccccgttc agtccgaccg
     2701 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gaaagacatg caaaagcacc
     2761 actggcagca gccactggta attgatttag aggagttagt cttgaagtca tgcgccggtt
     2821 aaggctaaac tgaaaggaca agttttggtg actgcgctcc tccaagccag ttacctcggt
     2881 tcaaagagtt ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt
     2941 tttcagagca agagattacg cgcagaccaa aacgatctca agaagatcat cttattaatc
     3001 agataaaata tttctagatt tcagtgcaat ttatctcttc aaatgtagca cctgaagtca
     3061 gccccatacg atataagttg ttactagtgc ttggattctc accaataaaa aacgcccggc
     3121 ggcaaccgag cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca
     3181 acaggagtcc aagcgagctc tcgaacccca gagtcccgct cagaagaact cgtcaagaag
     3241 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
     3301 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
     3361 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
     3421 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg
     3481 catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
     3541 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
     3601 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc
     3661 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc
     3721 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc
     3781 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cctgcagttc
     3841 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag
     3901 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag
     3961 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatgcgaaa
     4021 cgatcctcat cctgtctctt g
//
