LOCUS       sequence-244248-        4671 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Slot A, contains mKO2-MarA (step 1).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4671)
  AUTHORS   Santos-Moreno J, Schaerli Y
  TITLE     A Framework for the Modular and Combinatorial Assembly of Synthetic 
            Gene Circuits.
  JOURNAL   ACS Synth Biol. 2019 Jul 19;8(7):1691-1697. doi: 
            10.1021/acssynbio.9b00174. Epub 2019 Jun 24.
  PUBMED    31185158
REFERENCE   2  (bases 1 to 4671)
  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/acssynbio.9b00174"; journalName: "ACS Synth Biol"; date: 
            "2019-07-19- 19"; volume: "8"; issue: "7"; pages: "1691-1697"
COMMENT     Sequence Label: pTU-A-005
FEATURES             Location/Qualifiers
     source          1..4671
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    166..185
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     RBS             190..212
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             220..873
                     /codon_start=1
                     /product="monomeric Kusabira-Orange 2 fluorescent protein"
                     /label=mKO2
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     primer_bind     complement(1586..1604)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1600..1647
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1656..1742
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1834..1861
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             2364..2555
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2549..2571)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2657..2796
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2712..2731
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2811..2828)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2982..3570)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3062..3081)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3741..4532,4533..4601))
                     /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     4364..4383
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(4602..4671,1..35))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
       61 gaattcatag gatagattct ggaaacttta ccgtccgagc tcaggcttac cttactcgag
      121 caataaacag ttgatagggc ttctccgtta cagcctgcgg tccgggttca ctgccgtata
      181 ggcagtaatt ttgtttaact ttaagaagga gatatacata tggtttcggt tatcaaacca
      241 gagatgaaaa tgcgttacta tatggatggt tcagtaaatg gtcacgaatt tactattgag
      301 ggcgagggta cgggacgccc atacgagggg caccaggaaa tgactttacg cgtcacaatg
      361 gctgaaggcg ggcctatgcc gtttgcgttc gatcttgtta gtcatgtctt ttgttacggt
      421 caccgtgtat ttactaaata ccccgaggaa attccagact atttcaaaca agccttcccg
      481 gaaggtttgt cttgggagcg cagtttagag tttgaagacg gtggctcggc cagcgtgtca
      541 gctcatatta gtcttcgcgg caatacattt tatcacaagt caaagttcac cggcgtgaac
      601 ttccccgcag acggcccaat catgcagaat caaagtgttg attgggaacc gtccacagag
      661 aagattacag cttccgatgg agtcttaaag ggcgatgtaa ccatgtactt aaaattagaa
      721 gggggaggga accataaatg tcagatgaag actacctata aggccgcaaa agagattctt
      781 gaaatgcccg gagaccacta cattgggcat cgtttggtcc gtaagacaga aggaaatatt
      841 actgaacagg tcgaagacgc tgtggcacac agcatgtccc gccgtaatac tgacgccatc
      901 acaatccaca gcatcctgga ttggattgaa gacaacttgg agtcgccgtt gagtttagaa
      961 aaagttagtg aacgtagtgg ttactcaaag tggcaccttc agcgcatgtt taagaaggaa
     1021 acgggtcatt cattgggtca atatattcgt tctcgcaaga tgactgaaat tgcccagaaa
     1081 ttgaaagagt ctaatgaacc tattttgtac ctggcggagc gttacggctt tgaaagtcag
     1141 caaaccctta cacgtacctt caagaattac tttgacgttc caccacacaa atatcgtatg
     1201 accaacatgc agggtgagtc acgttttttg catccgttga atcattacaa ttcctaataa
     1261 tcgctgggac gcccgccatg gttcagccaa aaaacttaag accgccggtc ttgtccacta
     1321 ccttgcagta atgcggtgga caggatcggc ggttttcttt tctcttctca attcttctga
     1381 cctgtaacga ataatagata gtaaagtagt ctccgattga gttttctctg ccgagtccca
     1441 cccagttctg tgatttcagt aagttggtaa ttgatacact gttgcgagaa ctgctgcctg
     1501 gtagtagata ggttgttatt gagtaagaag gtaaagtgaa cgaaatccct gaaactgaga
     1561 ctgtagaaaa taagctttgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     1621 tctaaacggg tcttgagggg ttttttgcca ggcatcaaat aaaacgaaag gctcagtcgg
     1681 aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa
     1741 atccgccggg agcggatttg aacgttgtga agcaacggcc cggagggtgg cgggcaggac
     1801 gcccgccata aactgccagg catcaaacta agcagaaggc catcctgacg gatggccttt
     1861 ttgcgtttca gatctcgggc agcgttgggt cctggccacg ggtgcgcatg atcgtgctcc
     1921 tgtcgttgag gacccggcta ggctggcggg gttgccttac tggttagcag aatgaatcac
     1981 cgatacgcga gcgaacgtga agcgactgct gctgcaaaac gtctgcgacc tgagcaacaa
     2041 catgaatggt cttcggtttc cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct
     2101 gcaccattat gttccggatc tgcatcgcag gatgctgctg gctaccctgt ggaacaccta
     2161 catctgtatt aacgaagcgc tggcattgac cctgagtgat ttttctctgg tcccgccgca
     2221 tccataccgc cagttgttta ccctcacaac gttccagtaa ccgggcatgt tcatcatcag
     2281 taacccgtat cgtgagcatc ctctctcgtt tcatcggtat cattaccccc atgaacagaa
     2341 atccccctta cacggaggca tcagtgacca aacaggaaaa aaccgccctt aacatggccc
     2401 gctttatcag aagccagaca ttaacgcttc tggagaaact caacgagctg gacgcggatg
     2461 aacaggcaga catctgtgaa tcgcttcacg accacgctga tgagctttac cgcagctgcc
     2521 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     2581 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     2641 ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtatactg
     2701 gcttaactat gcggcatcag agcagattgt actgagagtg caccattgcg gtgtgaaata
     2761 ccgcacagat gcgtaaggag aaaataccgc atcaggcgct cttccgcttc ctcgctcact
     2821 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     2881 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     2941 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     3001 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3061 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3121 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     3181 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     3241 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     3301 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     3361 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     3421 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     3481 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     3541 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     3601 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     3661 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     3721 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     3781 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     3841 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     3901 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     3961 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     4021 ccagttaata gtttgcgcaa cgttgttgcc attgctgcag gcatcgtggt gtcacgctcg
     4081 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4141 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     4201 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     4261 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     4321 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaacac gggataatac cgcgccacat
     4381 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     4441 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     4501 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     4561 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     4621 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga a
//
