LOCUS       sequence-202216-        4331 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Template for T7 RNAP production of RNA transcript for expression of 
            firefly luciferase in a eukaryotic mammalian cell free lysate..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4331)
  AUTHORS   Canady TD, Telmer CA, Oyaghire SN, Armitage BA, Bruchez MP
  TITLE     In Vitro Reversible Translation Control Using gammaPNA Probes.
  JOURNAL   J Am Chem Soc. 2015 Aug 19;137(32):10268-75. doi: 
            10.1021/jacs.5b05351. Epub 2015 Aug 4.
  PUBMED    26241615
REFERENCE   2  (bases 1 to 4331)
  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: 
            "10.1021/jacs.5b05351"; journalName: "J Am Chem Soc"; date: 
            "2015-08-19- 19"; volume: "137"; issue: "32"; pages: "10268-75"
COMMENT     Sequence Label: T7-CMVtrans-FFLuc-polyA
FEATURES             Location/Qualifiers
     source          1..4331
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     15..32
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     113..132
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        113..131
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      174..183
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             180..1832
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(246..264)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     primer_bind     complement(1970..1989)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(2203..2222)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2382..2400)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2468..2572
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2573..2641,2642..3433)
                     /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(2791..2810)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3604..4192
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4093..4112
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac
       61 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcttatcgaa attaatacga
      121 ctcactatag ggagacccaa gctttcagat ccgctagcgc taccggggat ccagccacca
      181 tggaagacgc caaaaacata aagaaaggcc cggcgccatt ctatcctcta gaggatggaa
      241 ccgctggaga gcaactgcat aaggctatga agagatacgc cctggttcct ggaacaattg
      301 cttttacaga tgcacatatc gaggtgaaca tcacgtacgc ggaatacttc gaaatgtccg
      361 ttcggttggc agaagctatg aaacgatatg ggctgaatac aaatcacaga atcgtcgtat
      421 gcagtgaaaa ctctcttcaa ttctttatgc cggtgttggg cgcgttattt atcggagttg
      481 cagttgcgcc cgcgaacgac atttataatg aacgtgaatt gctcaacagt atgaacattt
      541 cgcagcctac cgtagtgttt gtttccaaaa aggggttgca aaaaattttg aacgtgcaaa
      601 aaaaattacc aataatccag aaaattatta tcatggattc taaaacggat taccagggat
      661 ttcagtcgat gtacacgttc gtcacatctc atctacctcc cggttttaat gaatacgatt
      721 ttgtaccaga gtcctttgat cgtgacaaaa caattgcact gataatgaat tcctctggat
      781 ctactgggtt acctaagggt gtggcccttc cgcatagaac tgcctgcgtc agattctcgc
      841 atgccagaga tcctattttt ggcaatcaaa tcattccgga tactgcgatt ttaagtgttg
      901 ttccattcca tcacggtttt ggaatgttta ctacactcgg atatttgata tgtggatttc
      961 gagtcgtctt aatgtataga tttgaagaag agctgttttt acgatccctt caggattaca
     1021 aaattcaaag tgcgttgcta gtaccaaccc tattttcatt cttcgccaaa agcactctga
     1081 ttgacaaata cgatttatct aatttacacg aaattgcttc tgggggcgca cctctttcga
     1141 aagaagtcgg ggaagcggtt gcaaaacgct tccatcttcc agggatacga caaggatatg
     1201 ggctcactga gactacatca gctattctga ttacacccga gggggatgat aaaccgggcg
     1261 cggtcggtaa agttgttcca ttttttgaag cgaaggttgt ggatctggat accgggaaaa
     1321 cgctgggcgt taatcagaga ggcgaattat gtgtcagagg acctatgatt atgtccggtt
     1381 atgtaaacaa tccggaagcg accaacgcct tgattgacaa ggatggatgg ctacattctg
     1441 gagacatagc ttactgggac gaagacgaac acttcttcat agttgaccgc ttgaagtctt
     1501 taattaaata caaaggatat caggtggccc ccgctgaatt ggaatcgata ttgttacaac
     1561 accccaacat cttcgacgcg ggcgtggcag gtcttcccga cgatgacgcc ggtgaacttc
     1621 ccgccgccgt tgttgttttg gagcacggaa agacgatgac ggaaaaagag atcgtggatt
     1681 acgtcgccag tcaagtaaca accgcgaaaa agttgcgcgg aggagttgtg tttgtggacg
     1741 aagtaccgaa aggtcttacc ggaaaactcg acgcaagaaa aatcagagag atcctcataa
     1801 aggccaagaa gggcggaaag tccaaattgt aaaatgtaac tgtattcagc gatgacgaaa
     1861 ttcttagcta ttgtaatcct ccgagggggc gagctcccaa aaaaaaaaaa aaaaaaaaaa
     1921 aaaaaaaacc gaattggcat caccggcgcc acaggtgcgg ttgctggcgc ctatatcgcc
     1981 gacatcaccg atggggaaga tcgggctcgc cacttcgggc tcatgagcgc ttgtttcggc
     2041 gtgggtatgg tggcaggccc cgtggccggg ggactgttgg gcgccatctc cttgcatgca
     2101 ccattccttg cggcggcggt gctcaacggc ctcaacctac tactgggctg cttcctaatg
     2161 caggagtcgc ataagggaga gcgtcgaatg gtgcactctc agtacaatct gctctgatgc
     2221 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     2281 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     2341 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     2401 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     2461 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     2521 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     2581 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     2641 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     2701 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     2761 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     2821 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     2881 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     2941 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     3001 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     3061 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     3121 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     3181 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     3241 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     3301 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     3361 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     3421 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     3481 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     3541 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     3601 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     3661 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     3721 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     3781 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     3841 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     3901 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     3961 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     4021 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4081 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4141 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4201 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     4261 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     4321 tcgccgcagc c
//
