LOCUS       sequence-370380-        4568 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Firefly luciferase via a T7 promoter in TXTL.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4568)
  AUTHORS   Sato W, Rasmussen M, Deich C, Engelhart AE, Adamala KP
  TITLE     Expanding luciferase reporter systems for cell-free protein 
            expression.
  JOURNAL   Sci Rep. 2022 Jul 7;12(1):11489. doi: 10.1038/s41598-022-15624-6.
  PUBMED    35798760
REFERENCE   2  (bases 1 to 4568)
  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 Rep."; 
            date: "2022-07-7"; volume: "12(1):11489. doi"; pages: " 
            10.1038/s41598-022-15624-6"
COMMENT     Sequence Label: pTXTL-T7max-FLuc
FEATURES             Location/Qualifiers
     source          1..4568
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     36..55
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        36..54
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             63..85
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             104..1753
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(170..188)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     CDS             1769..1792
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     rep_origin      2014..2469
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2101..2120)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2311..2332
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2536..2555)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2655..2677
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2715..2733)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2801..2905
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2906..2974,2975..3766)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3124..3143)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3937..4525
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4426..4445
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggccttttgc tcacatggct cgacagatct aattctaata cgactcacta tagggaaata
       61 attttgttta actttaagaa ggagatatac cggtatgact agtatggaag acgccaaaaa
      121 cataaagaaa ggcccggcgc cattctatcc gctggaagat ggaaccgctg gagagcaact
      181 gcataaggct atgaagagat acgccctggt tcctggaaca attgctttta cagatgcaca
      241 tatcgaggtg gacatcactt acgctgagta cttcgaaatg tccgttcggt tggcagaagc
      301 tatgaaacga tatgggctga atacaaatca cagaatcgtc gtatgcagtg aaaactctct
      361 tcaattcttt atgccggtgt tgggcgcgtt atttatcgga gttgcagttg cgcccgcgaa
      421 cgacatttat aatgaacgtg aattgctcaa cagtatgggc atttcgcagc ctaccgtggt
      481 gttcgtttcc aaaaaggggt tgcaaaaaat tttgaacgtg caaaaaaagc tcccaatcat
      541 ccaaaaaatt attatcatgg attctaaaac ggattaccag ggatttcagt cgatgtacac
      601 gttcgtcaca tctcatctac ctcccggttt taatgaatac gattttgtgc cagagtcctt
      661 cgatagggac aagacaattg cactgatcat gaactcctct ggatctactg gtctgcctaa
      721 aggtgtcgct ctgcctcata gaactgcctg cgtgagattc tcgcatgcca gagatcctat
      781 ttttggcaat caaatcattc cggatactgc gattttaagt gttgttccat tccatcacgg
      841 ttttggaatg tttactacac tcggatattt gatatgtgga tttcgagtcg tcttaatgta
      901 tagatttgaa gaagagctgt ttctgaggag ccttcaggat tacaagattc aaagtgcgct
      961 gctggtgcca accctattct ccttcttcgc caaaagcact ctgattgaca aatacgattt
     1021 atctaattta cacgaaattg cttctggtgg cgctcccctc tctaaggaag tcggggaagc
     1081 ggttgccaag aggttccatc tgccaggtat caggcaagga tatgggctca ctgagactac
     1141 atcagctatt ctgattacac ccgaggggga tgataaaccg ggcgcggtcg gtaaagttgt
     1201 tccatttttt gaagcgaagg ttgtggatct ggataccggg aaaacgctgg gcgttaatca
     1261 aagaggcgaa ctgtgtgtga gaggtcctat gattatgtcc ggttatgtaa acaatccgga
     1321 agcgaccaac gccttgattg acaaggatgg atggctacat tctggagaca tagcttactg
     1381 ggacgaagac gaacacttct tcatcgttga ccgcctgaag tctctgatta agtacaaagg
     1441 ctatcaggtg gctcccgctg aattggaatc catcttgctc caacacccca acatcttcga
     1501 cgcaggtgtc gcaggtcttc ccgacgatga cgccggtgaa cttcccgccg ccgttgttgt
     1561 tttggagcac ggaaagacga tgacggaaaa agagatcgtg gattacgtcg ccagtcaagt
     1621 aacaaccgcg aaaaagttgc gcggaggagt tgtgtttgtg gacgaagtac cgaaaggtct
     1681 taccggaaaa ctcgacgcaa gaaaaatcag agagatcctc ataaaggcca agaagggcgg
     1741 aaagatcgcc gtgacgcgtg gaggtggcca tcaccatcat caccatcacc acggataact
     1801 cgagcaaagc ccgccgaaag gcgggctttt ctgtacaaca acaattgcat tcattttatg
     1861 tttcaggttc agggggagat gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt
     1921 ggtaaaatcg ataaggatcc gggctggcgt aatagcgaag aggcccgcac cgatcgccct
     1981 tcccaacagt tgcgcagcct gaatggcgaa tggacgcgcc ctgtagcggc gcattaagcg
     2041 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     2101 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     2161 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     2221 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     2281 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     2341 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     2401 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     2461 ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     2521 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     2581 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     2641 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     2701 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     2761 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     2821 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     2881 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     2941 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     3001 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     3061 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     3121 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     3181 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     3241 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     3301 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     3361 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     3421 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     3481 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     3541 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     3601 atctggagcc ggtgagcgtg ggtcgcgcgg tatcattgca gcactggggc cagatggtaa
     3661 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     3721 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     3781 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     3841 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3901 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3961 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     4021 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     4081 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     4141 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     4201 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     4261 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     4321 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     4381 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     4441 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     4501 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4561 cttttgct
//
