LOCUS       sequence-417971-        5632 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  nonsense mistranslation, TGA stop variant 2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5632)
  AUTHORS   Hinnu M, Putrins M, Kogermann K, Kaldalu N, Tenson T
  TITLE     Fluorescent reporters give new insights into antibiotics-induced 
            nonsense and frameshift mistranslation.
  JOURNAL   Sci Rep. 2024 Mar 22;14(1):6883. doi: 10.1038/s41598-024-57597-8.
  PUBMED    38519558
REFERENCE   2  (bases 1 to 5632)
  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: "2024-03-22"; volume: "14(1):6883. doi"; pages: " 
            10.1038/s41598-024-57597-8"
COMMENT     Sequence Label: pSC101-GFPmut2-mScarlet-I TGATC
FEATURES             Location/Qualifiers
     source          1..5632
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     118..137
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(join(196..987,988..1056))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     819..838
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1057..1161)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(1213..1240)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        join(1448..1453,1454..1470,1471..1476)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1576..1604)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2199..2219
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             2280..2981
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKG*SEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQ
                     TAKLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFED
                     GGAVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGD
                     IKMALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEG
                     RHSTGGMDELYK"
     primer_bind     complement(2419..2437)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2687..2706
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3084..3107
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     terminator      3192..3286
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      3778..4000
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             4048..4998
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     terminator      complement(5536..5622)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 agcttgcatg cccaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
       61 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
      121 gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat
      181 gcgtccggcg tagagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
      241 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
      301 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
      361 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
      421 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
      481 taatagtttg cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac gctcgtcgtt
      541 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
      601 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
      661 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
      721 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
      781 gcggcgaccg agttgctctt gcccggcgtc aacacgggat aataccgcgc cacatagcag
      841 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
      901 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
      961 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     1021 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     1081 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     1141 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     1201 cattattatc ataaaaggcc atccgtcagg atggccttct atgaatgcct tggctgcgat
     1261 tcattcttta tatgaataaa attgttgtca gtttcacgtc ttgtcctgcc atatcgcgaa
     1321 atttctgcgc aaaagcacaa aaaatttttg catctccccc ttgatgacgt gggttacgac
     1381 cccatttagt agtcaaccgc agtgagtgag tctgacgagg ccctttcgtc ttcaagaatt
     1441 ctcatgtttg acagcttatc atcgataagc tttaatgcgg tagtttatca cagttaaatt
     1501 gctaacgcag tccggggaat tcaggcttgg aggaacaata tgaaaggatc caaaggtgaa
     1561 gaattattca ctggagttgt cccaattctt gttgaattag atggtgatgt taatgggcac
     1621 aaattttctg tcagtggaga gggtgaaggt gatgcaacat acggaaaact tacccttaaa
     1681 tttatttgca ctactggaaa actacctgtt ccatggccaa cacttgtcac tactttcgcg
     1741 tatggtcttc aatgctttgc gagataccca gatcatatga aacagcatga ctttttcaag
     1801 agtgccatgc ccgaaggtta tgtacaggaa agaactatat ttttcaaaga tgacgggaac
     1861 tacaagacac gtgctgaagt caagtttgaa ggtgataccc ttgttaatag aatcgagtta
     1921 aaaggtattg attttaaaga agatggaaac attcttggac acaaattgga atacaactat
     1981 aactcacaca aagtatacat catggcagac aaacaaaagg atggaatcaa agttaacttc
     2041 aaaattagac acaacattga agatggaagc gttcaactag cagaccatta tcaacgaaat
     2101 actccaattg gcgatggccc tgtcctttta ccagacaacc attacctgtc cacacaatct
     2161 gccctttcga aagatcccaa cgaaaagaga gaccacatgg tccttcttga gtttgtaaca
     2221 gctgctggga ttacacatgg tatggatgaa ttgtacaaat aaccgccagg aggaacaata
     2281 tggtgagcaa gggctgatct gaggcagtga tcaaggagtt catgcggttc aaggtgcaca
     2341 tggagggctc catgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     2401 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttctcct
     2461 gggacatcct gtcccctcag ttcatgtacg gctccagggc cttcatcaag caccccgccg
     2521 acatccccga ctactataag cagtccttcc ccgagggctt caagtgggag cgcgtgatga
     2581 acttcgagga cggcggcgcc gtgaccgtga cccaggacac ctccctggag gacggcaccc
     2641 tgatctacaa ggtgaagctc cgcggcacca acttccctcc tgacggcccc gtaatgcaga
     2701 agaagacaat gggctgggaa gcgtccaccg agcggttgta ccccgaggac ggcgtgctga
     2761 agggcgacat taagatggcc ctgcgcctga aggacggcgg ccgctacctg gcggacttca
     2821 agaccaccta caaggccaag aagcccgtgc agatgcccgg cgcctacaac gtcgaccgca
     2881 agttggacat cacctcccac aacgaggact acaccgtggt ggaacagtac gaacgctccg
     2941 agggccgcca ctccaccggc ggcatggacg agctgtacaa gtgaccgcct aatgagcggg
     3001 ctttaatcac tagtgaattc gcggccgcct gcaggtcgac catatgggag agctccgtcg
     3061 acaagcttgc ggccgcactc gagcaccacc accaccacca ccaccactaa ttgattaata
     3121 cctaggctgc taaacaaagc ccatggatcg atagctggtc gacaggactc tggggttcga
     3181 gagctcgctt ggactcctgt tgatagatcc agtaatgacc tcagaactcc atctggattt
     3241 gttcagaacg ctcggttgcc gccgggcgtt ttttattggt gagaatccaa gcactaggga
     3301 cagtaagacg ggtaagcctg ttgatgatac cgctgcctta ctgggtgcat tagccagtct
     3361 gaatgacctg tcacgggata atccgaagtg gtcagactgg aaaatcagag ggcaggaact
     3421 gctgaacagc aaaaagtcag atagcaccac atagcagacc cgccataaaa cgccctgaga
     3481 agcccgtgac gggcttttct tgtattatgg gtagtttcct tgcatgaatc cataaaaggc
     3541 gcctgtagtg ccatttaccc ccattcactg ccagagccgt gagcgcagcg aactgaatgt
     3601 cacgaaaaag acagcgactc aggtgcctga tggtcggaga caaaaggaat attcagcgat
     3661 ttgcccgagc ttgcgagggt gctacttaag cctttagggt tttaaggtct gttttgtaga
     3721 ggagcaaaca gcgtttgcga catccttttg taatactgcg gaactgacta aagtagtgag
     3781 ttatacacag ggctgggatc tattcttttt atcttttttt attctttctt tattctataa
     3841 attataacca cttgaatata aacaaaaaaa acacacaaag gtctagcgga atttacagag
     3901 ggtctagcag aatttacaag ttttccagca aaggtctagc agaatttaca gatacccaca
     3961 actcaaagga aaaggactag taattatcat tgactagccc atctcaattg gtatagtgat
     4021 taaaatcacc tagaccaatt gagatgtatg tctgaattag ttgttttcaa agcaaatgaa
     4081 ctagcgatta gtcgctatga cttaacggag catgaaacca agctaatttt atgctgtgtg
     4141 gcactactca accccacgat tgaaaaccct acaaggaaag aacggacggt atcgttcact
     4201 tataaccaat acgctcagat gatgaacatc agtagggaaa atgcttatgg tgtattagct
     4261 aaagcaacca gagagctgat gacgagaact gtggaaatca ggaatccttt ggttaaaggc
     4321 tttgagattt tccagtggac aaactatgcc aagttctcaa gcgaaaaatt agaattagtt
     4381 tttagtgaag agatattgcc ttatcttttc cagttaaaaa aattcataaa atataatctg
     4441 gaacatgtta agtcttttga aaacaaatac tctatgagga tttatgagtg gttattaaaa
     4501 gaactaacac aaaagaaaac tcacaaggca aatatagaga ttagccttga tgaatttaag
     4561 ttcatgttaa tgcttgaaaa taactaccat gagtttaaaa ggcttaacca atgggttttg
     4621 aaaccaataa gtaaagattt aaacacttac agcaatatga aattggtggt tgataagcga
     4681 ggccgcccga ctgatacgtt gattttccaa gttgaactag atagacaaat ggatctcgta
     4741 accgaacttg agaacaacca gataaaaatg aatggtgaca aaataccaac aaccattaca
     4801 tcagattcct acctacataa cggactaaga aaaacactac acgatgcttt aactgcaaaa
     4861 attcagctca ccagttttga ggcaaaattt ttgagtgaca tgcaaagtaa gtatgatctc
     4921 aatggttcgt tctcatggct cacgcaaaaa caacgaacca cactagagaa catactggct
     4981 aaatacggaa ggatctgagg ttcttatggc tcttgtatct atcagtgaag catcaagact
     5041 aacaaacaaa agtagaacaa ctgttcaccg ttacatatca aagggaaaac tgtccatatg
     5101 cacagatgaa aacggtgtaa aaaagataga tacatcagag cttttacgag tttttggtgc
     5161 attcaaagct gttcaccatg aacagatcga caatgtaaca gatgaacagc atgtaacacc
     5221 taatagaaca ggtgaaacca gtaaaacaaa gcaactagaa catgaaattg aacacctgag
     5281 acaacttgtt acagctcaac agtcacacat agacagcctg aaacaggcga tgctgcttat
     5341 cgaatcaaag ctgccgacaa cacgggagcc agtgacgcct cccgtgggga aaaaatcatg
     5401 gcaattctgg aagaaatagc gctttcagcc ggcaaaccgg ctgaagccgg atctgcgatt
     5461 ctgataacaa actagcaaca ccagaacagc ccgtttgcgg gcagcaaaac ccgtacccta
     5521 ggtctagggc ggcggatttg tcctactcag gagagcgttc accgacaaac aacagataaa
     5581 acgaaaggcc cagtctttcg actgagcctt tcgttttatt tgatgcctct ag
//
