LOCUS       sequence-454080-        7148 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1, Position 2 containing 35S::CUP2-p65::NOS..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7148)
  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 7148)
  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: pCHW101020
FEATURES             Location/Qualifiers
     source          1..7148
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    42..66
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(222..1370)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      complement(1706..2294)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1786..1805)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2465..3256,3257..3325))
                     /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     3088..3107
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3326..3430)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3517..4148
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    4240..4264
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     promoter        5362..5707
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     5650..5670
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             6482..6853
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALLSQISS"
     CDS             6854..6874
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      6890..7142
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
ORIGIN
        1 actattgtct tctgcacgaa gtggtttaaa ctatcagtgt ttgacaggat atattggcgg
       61 gtaaacctaa gagaaaagag cgtttattag aataatcgga tatttaaaag ggcgtgaaaa
      121 ggtttatccg ttcgtccatt tgtatgtgca tgccaaccac agggttcccc agatcaggcg
      181 ctggctgctg aacccccagc cggaactgac cccacaaggc cctagcgttt gcaatgcacc
      241 aggtcatcat tgacccaggc gtgttccacc aggccgctgc ctcgcaactc ttcgcaggct
      301 tcgccgacct gctcgcgcca cttcttcacg cgggtggaat ccgatccgca catgaggcgg
      361 aaggtttcca gcttgagcgg gtacggctcc cggtgcgagc tgaaatagtc gaacatccgt
      421 cgggccgtcg gcgacagctt gcggtacttc tcccatatga atttcgtgta gtggtcgcca
      481 gcaaacagca cgacgatttc ctcgtcgatc aggacctggc aacgggacgt tttcttgcca
      541 cggtccagga cgcggaagcg gtgcagcagc gacaccgatt ccaggtgccc aacgcggtcg
      601 gacgtgaagc ccatcgccgt cgcctgtagg cgcgacaggc attcctcggc cttcgtgtaa
      661 taccggccat tgatcgacca gcccaggtcc tggcaaagct cgtagaacgt gaaggtgatc
      721 ggctcgccga taggggtgcg cttcgcgtac tccaacacct gctgccacac cagttcgtca
      781 tcgtcggccc gcagctcgac gccggtgtag gtgatcttca cgtccttgtt gacgtggaaa
      841 atgaccttgt tttgcagcgc ctcgcgcggg attttcttgt tgcgcgtggt gaacagggca
      901 gagcgggccg tgtcgtttgg catcgctcgc atcgtgtccg gccacggcgc aatatcgaac
      961 aaggaaagct gcatttcctt gatctgctgc ttcgtgtgtt tcagcaacgc ggcctgcttg
     1021 gcctcgctga cctgttttgc caggtcctcg ccggcggttt ttcgcttctt ggtcgtcata
     1081 gttcctcgcg tgtcgatggt catcgacttc gccaaacctg ccgcctcctg ttcaagacga
     1141 cgcgaacgct ccacggcggc cgatggcgcg ggcagggcag ggggagccag ttgcacgctg
     1201 tcgcgctcga tcttggccgt agcttgctgg accatcgagc cgacggactg gaaggtttcg
     1261 cggggcgcac gcatgacggt gcggcttgcg atggtttcgg catcctcggc ggaaaacccc
     1321 gcgtcgatca gttcttgcct gtatgccttc cggtcaaacg tccgattcat tcaccctcct
     1381 tgcgggattg ccccgactca cgccggggca atgtgccctt attcctgatt tgacccgcct
     1441 ggtgccttgg tgtccagata atccacctta tcggcaatga agtcggtccc gtagaccgtc
     1501 tggccgtcct tctcgtactt ggtattccga atcttgccct gcacgaatac cagcgacccc
     1561 ttgcccaaat acttgccgtg ggcctcggcc tgagagccaa aacacttgat gcggaagaag
     1621 tcggtgcgct cctgcttgtc gccggcatcg ttgcgccaca tctaggatct gccaggaacc
     1681 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     1741 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     1801 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     1861 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     1921 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     1981 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     2041 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     2101 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     2161 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     2221 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     2281 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     2341 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     2401 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     2461 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     2521 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     2581 gccccagtgc tgcaatgata ccgcgagaac cacgctcacc ggctccagat ttatcagcaa
     2641 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     2701 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     2761 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     2821 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     2881 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     2941 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     3001 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     3061 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     3121 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     3181 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     3241 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     3301 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     3361 aaggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     3421 gggttccgcg cacgaattgg ccagcgctgc catttttggg gtgaggccgt tcgcggccga
     3481 ggggcgcagc ccctgggggg atgggaggcc cgcgttagcg ggccgggagg gttcgagaag
     3541 ggggggcacc ccccttcggc gtgcgcggtc acgcgcacag ggcgcagccc tggttaaaaa
     3601 caaggtttat aaatattggt ttaaaagcag gttaaaagac aggttagcgg tggccgaaaa
     3661 acgggcggaa acccttgcaa atgctggatt ttctgcctgt ggacagcccc tcaaatgtca
     3721 ataggtgcgc ccctcatctg tcagcactct gcccctcaag tgtcaaggat cgcgcccctc
     3781 atctgtcagt agtcgcgccc ctcaagtgtc aataccgcag ggcacttatc cccaggcttg
     3841 tccacatcat ctgtgggaaa ctcgcgtaaa atcaggcgtt ttcgccgatt tgcgaggctg
     3901 gccagctcca cgtcgccggc cgaaatcgag cctgcccctc atctgtcaac gccgcgccgg
     3961 gtgagtcggc ccctcaagtg tcaacgtccg cccctcatct gtcagtgagg gccaagtttt
     4021 ccgcgaggta tccacaacgc cggcggccgc ggtgtctcgc acacggcttc gacggcgttt
     4081 ctggcgcgtt tgcagggcca tagacggccg ccagcccagc ggcgagggca accagcccgg
     4141 tgagcgtcgc aaaggagatc ctgatctgac tgatgggctg cctgtatcga gtggtgattt
     4201 tgtgccgagc tgccggtcgg ggagctgttg gctggctggt ggcaggatat attgtggtgt
     4261 aaacaaattg acgcttagac aacttaataa cacattgcgg acgtttttaa tgtactgggg
     4321 tggatgcagt gggccccact ctgtgaagac aagcaagaat tcaagcttgg aggaattcca
     4381 atcccacaaa aatctgagct taacagcaca gttgctcctc tcagagcaga atcgggtatt
     4441 caacaccctc atatcaacta ctacgttgtg tataacggtc cacatgccgg tatatacgat
     4501 gactggggtt gtacaaaggc ggcaacaaac ggcgttcccg gagttgcaca caagaaattt
     4561 gccactatta cagaggcaag agcagcagct gacgcgtaca caacaagtca gcaaacagac
     4621 aggttgaact tcatccccaa aggagaagct caactcaagc ccaagagctt tgctaaggcc
     4681 ctaacaagcc caccaaagca aaaagcccac tggctcacgc taggaaccaa aaggcccagc
     4741 agtgatccag ccccaaaaga gatctccttt gccccggaga ttacaatgga cgatttcctc
     4801 tatctttacg atctaggaag gaagttcgaa ggtgaaggtg acgacactat gttcaccact
     4861 gataatgaga aggttagcct cttcaatttc agaaagaatg ctgacccaca gatggttaga
     4921 gaggcctacg cagcaagtct catcaagacg atctacccga gtaacaatct ccaggagatc
     4981 aaataccttc ccaagaaggt taaagatgca gtcaaaagat tcaggactaa ttgcatcaag
     5041 aacacagaga aagacatatt tctcaagatc agaagtacta ttccagtatg gacgattcaa
     5101 ggcttgcttc ataaaccaag gcaagtaata gagattggag tctctaaaaa ggtagttcct
     5161 actgaatcta aggccatgca tggagtctaa gattcaaatc gaggatctaa cagaactcgc
     5221 cgtcaagact ggcgaacagt tcatacagag tcttttacga ctcaatgaca agaagaaaat
     5281 cttcgtcaac atggtggagc acgacactct ggtctactcc aaaaatgtca aagatacagt
     5341 ctcagaagat caaagggcta ttgagacttt tcaacaaagg ataatttcgg gaaacctcct
     5401 cggattccat tgcccagcta tctgtcactt catcgaaagg acagtagaaa aggaaggtgg
     5461 ctcctacaaa tgccatcatt gcgataaagg aaaggctatc attcaagatc tctctgccga
     5521 cagtggtccc aaagatggac ccccacccac gaggagcatc gtggaaaaag aagaggttcc
     5581 aaccacgtct acaaagcaag tggattgatg tgacatctcc actgacgtaa gggatgacgc
     5641 acaatcccac tatccttcgc aagacccttc ctctatataa ggaagttcat ttcatttgga
     5701 gaggacacgc tcgagtataa gagctcattt ttacaacaat taccaacaac aacaaacaac
     5761 aaacaacatt acaattacat ttacaattat cgatacaatg gtcgtaatta acggggtcaa
     5821 atatgcctgt gaaacgtgta tcaggggtca cagggcggcg cagtgtactc acactgatgg
     5881 tccgctacag atgatcagac gcaagggtag accatcgacc acatgtggcc attgtaaaga
     5941 gctgagaaga accaagaact tcaacccatc cggtgggtgc atgtgtgcct ctgcacgacg
     6001 gccagctgtt ggcagcaagg aagatgaaac acgatgtcgt tgtgatgagg gtgaaccttg
     6061 taaatgtcat accaagagga aaagcagccg gaaatcaaag ggagggtcat gccacagaag
     6121 ggcaaatgat gaagcagcgc atgtcaatgg tcttggtatt gcagatctgg acgttctttt
     6181 gggcctaaat ggtcgctcgt cggatgtaga catgacaacc acattgccga gtttgaagcc
     6241 acctctgcaa aacggagaaa ttaaggccga cagcattgac aatcttgatt tggcttccct
     6301 cgatccgctt gagcaaagcc ctagtatatc tatggaacct gttagtatca atgaaacagg
     6361 aagcgcatat acaactacga acacagcact aaacgatatt gacattccat tctccatcaa
     6421 tgagttgaac gagctataca aacaagtatc ttcgcataac tcacattcac agccatattc
     6481 gccaacgcag gcgggcgaag gaaccctgag cgaggccctc ttgcagcttc agttcgatga
     6541 cgaagatttg ggagccctgc tgggtaacag cacagaccct gccgtattca ccgatctcgc
     6601 atccgtggac aacagcgagt ttcagcagct cttgaatcag ggaatcccgg tcgcacctca
     6661 taccacagag cccatgctga tggaataccc ggaggctatc acgcgactgg tgaccggcgc
     6721 acagcgacca cccgatccag cccctgcccc actgggtgcc ccgggtttgc ccaatggcct
     6781 cctcagcggc gatgaggatt tctccagcat cgctgatatg gatttctccg ctttgctgag
     6841 ccagataagc tcccctaaaa agaagcgtaa agtctaagct tgtcaagcag atcgttcaaa
     6901 catttggcaa taaagtttct taagattgaa tcctgttgcc ggtcttgcga tgattatcat
     6961 ataatttctg ttgaattacg ttaagcatgt aataattaac atgtaatgca tgacgttatt
     7021 tatgagatgg gtttttatga ttagagtccc gcaattatac atttaatacg cgatagaaaa
     7081 caaaatatag cgcgcaaact aggataaatt atcgcgcgcg gtgtcatcta tgttactaga
     7141 tcgacgct
//
