LOCUS       sequence-454091-        5552 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 acceptor, Position 3 containing CBS4-miniDFR::LacZ::nosT..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5552)
  AUTHORS   Chiang BJ, Lin KY, Chen YF, Huang CY, Goh FJ, Huang LT, Chen LH, Wu 
            CH
  TITLE     Development of a tightly regulated copper-inducible transient gene 
            expression system in Nicotiana benthamiana incorporating suicide 
            exon and Cre recombinase
  JOURNAL   bioRxiv 2024.03.23.586378
REFERENCE   2  (bases 1 to 5552)
  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: "bioRxiv 
            2024.03.23.586378"
COMMENT     Sequence Label: pCHW111111
FEATURES             Location/Qualifiers
     source          1..5552
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(36..624)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(116..135)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(795..1586,1587..1655))
                     /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     1418..1437
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1656..1760)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1847..2478
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    2570..2594
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     protein_bind    3101..3122
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3137..3142,3143..3160,3161..3167)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3175..3191
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     3180..3202
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3199..3215
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3199..3215
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             3211..3534
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    3228..3284
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(3285..3302)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3285..3301)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3294..3316)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3510..3529)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     terminator      3624..3876
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    3924..3948
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(4104..5252)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
ORIGIN
        1 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
       61 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
      121 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
      181 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
      241 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
      301 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
      361 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
      421 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
      481 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
      541 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
      601 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
      661 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
      721 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
      781 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
      841 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
      901 ttaccatctg gccccagtgc tgcaatgata ccgcgagaac cacgctcacc ggctccagat
      961 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     1021 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     1081 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     1141 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     1201 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     1261 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     1321 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     1381 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     1441 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     1501 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     1561 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     1621 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     1681 agcatttatc aaggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     1741 aaacaaatag gggttccgcg cacgaattgg ccagcgctgc catttttggg gtgaggccgt
     1801 tcgcggccga ggggcgcagc ccctgggggg atgggaggcc cgcgttagcg ggccgggagg
     1861 gttcgagaag ggggggcacc ccccttcggc gtgcgcggtc acgcgcacag ggcgcagccc
     1921 tggttaaaaa caaggtttat aaatattggt ttaaaagcag gttaaaagac aggttagcgg
     1981 tggccgaaaa acgggcggaa acccttgcaa atgctggatt ttctgcctgt ggacagcccc
     2041 tcaaatgtca ataggtgcgc ccctcatctg tcagcactct gcccctcaag tgtcaaggat
     2101 cgcgcccctc atctgtcagt agtcgcgccc ctcaagtgtc aataccgcag ggcacttatc
     2161 cccaggcttg tccacatcat ctgtgggaaa ctcgcgtaaa atcaggcgtt ttcgccgatt
     2221 tgcgaggctg gccagctcca cgtcgccggc cgaaatcgag cctgcccctc atctgtcaac
     2281 gccgcgccgg gtgagtcggc ccctcaagtg tcaacgtccg cccctcatct gtcagtgagg
     2341 gccaagtttt ccgcgatgta tccacaacgc cggcggccgc ggtgtctcgc acacggcttc
     2401 gacggcgttt ctggcgcgtt tgcagggcca tagacggccg ccagcccagc ggcgagggca
     2461 accagcccgg tgagcgtcgc aaaggagatc ctgatctgac tgatgggctg cctgtatcga
     2521 gtggtgattt tgtgccgagc tgccggtcgg ggagctgttg gctggctggt ggcaggatat
     2581 attgtggtgt aaacaaattg acgcttagac aacttaataa cacattgcgg acgtttttaa
     2641 tgtactgggg tggatgcagt gggccccact ctgtgaagac aaactagaat tcgagctcgg
     2701 agagtggtct tgggagagcg atgcgtcttt tccgatgaac cgttccagca aaaaagacta
     2761 agcgatgcgt cttttccgct gaaccgttcc agcaaaaaag actaagcgat gcgtcttttc
     2821 cgctgaaccg ttccagcaaa aaagactagg agagcgatgc gtcttttccg ctgaaccgtt
     2881 ccagcaaaaa agactatccc gtcctaatac ctctacgtgc aagcagtata aaaacaaaat
     2941 acataaacat aactaaaatc tcatatcgtt gtaattgtca tatactagtg gtaattaagt
     3001 ccagttacac ttaaggttaa atttgctgac tcttttttac ctttcatttc tgaaaaatgt
     3061 gagaccccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     3121 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     3181 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag cttgcatgcc
     3241 tgcaggtcga ctctagagga tccccgggta ccgagctcga attcactggc cgtcgtttta
     3301 caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc
     3361 cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     3421 cgcagcctga atggcgaatg gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     3481 atttcacacc gcatatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     3541 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     3601 tccgggtctc agcttgtcaa gcagatcgtt caaacatttg gcaataaagt ttcttaagat
     3661 tgaatcctgt tgccggtctt gcgatgatta tcatataatt tctgttgaat tacgttaagc
     3721 atgtaataat taacatgtaa tgcatgacgt tatttatgag atgggttttt atgattagag
     3781 tcccgcaatt atacatttaa tacgcgatag aaaacaaaat atagcgcgca aactaggata
     3841 aattatcgcg cgcggtgtca tctatgttac tagatcgacg ctttacttgt cttctgcacg
     3901 aagtggttta aactatcagt gtttgacagg atatattggc gggtaaacct aagagaaaag
     3961 agcgtttatt agaataatcg gatatttaaa agggcgtgaa aaggtttatc cgttcgtcca
     4021 tttgtatgtg catgccaacc acagggttcc ccagatcagg cgctggctgc tgaaccccca
     4081 gccggaactg accccacaag gccctagcgt ttgcaatgca ccaggtcatc attgacccag
     4141 gcgtgttcca ccaggccgct gcctcgcaac tcttcgcagg cttcgccgac ctgctcgcgc
     4201 cacttcttca cgcgggtgga atccgatccg cacatgaggc ggaaggtttc cagcttgagc
     4261 gggtacggct cccggtgcga gctgaaatag tcgaacatcc gtcgggccgt cggcgacagc
     4321 ttgcggtact tctcccatat gaatttcgtg tagtggtcgc cagcaaacag cacgacgatt
     4381 tcctcgtcga tcaggacctg gcaacgggac gttttcttgc cacggtccag gacgcggaag
     4441 cggtgcagca gcgacaccga ttccaggtgc ccaacgcggt cggacgtgaa gcccatcgcc
     4501 gtcgcctgta ggcgcgacag gcattcctcg gccttcgtgt aataccggcc attgatcgac
     4561 cagcccaggt cctggcaaag ctcgtagaac gtgaaggtga tcggctcgcc gataggggtg
     4621 cgcttcgcgt actccaacac ctgctgccac accagttcgt catcgtcggc ccgcagctcg
     4681 acgccggtgt aggtgatctt cacgtccttg ttgacgtgga aaatgacctt gttttgcagc
     4741 gcctcgcgcg ggattttctt gttgcgcgtg gtgaacaggg cagagcgggc cgtgtcgttt
     4801 ggcatcgctc gcatcgtgtc cggccacggc gcaatatcga acaaggaaag ctgcatttcc
     4861 ttgatctgct gcttcgtgtg tttcagcaac gcggcctgct tggcctcgct gacctgtttt
     4921 gccaggtcct cgccggcggt ttttcgcttc ttggtcgtca tagttcctcg cgtgtcgatg
     4981 gtcatcgact tcgccaaacc tgccgcctcc tgttcaagac gacgcgaacg ctccacggcg
     5041 gccgatggcg cgggcagggc agggggagcc agttgcacgc tgtcgcgctc gatcttggcc
     5101 gtagcttgct ggaccatcga gccgacggac tggaaggttt cgcggggcgc acgcatgacg
     5161 gtgcggcttg cgatggtttc ggcatcctcg gcggaaaacc ccgcgtcgat cagttcttgc
     5221 ctgtatgcct tccggtcaaa cgtccgattc attcaccctc cttgcgggat tgccccgact
     5281 cacgccgggg caatgtgccc ttattcctga tttgacccgc ctggtgcctt ggtgtccaga
     5341 taatccacct tatcggcaat gaagtcggtc ccgtagaccg tctggccgtc cttctcgtac
     5401 ttggtattcc gaatcttgcc ctgcacgaat accagcgacc ccttgcccaa atacttgccg
     5461 tgggcctcgg cctgagagcc aaaacacttg atgcggaaga agtcggtgcg ctcctgcttg
     5521 tcgccggcat cgttgcgcca catctaggat ct
//
