LOCUS       sequence-378261-        8847 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To control for variation introduced to the gene circuits, Rluc 
            expression not affected by the introduction of terminators or 
            recombination target sites..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8847)
  AUTHORS   Lloyd JPB, Ly F, Gong P, Pflueger J, Swain T, Pflueger C, Fourie E, 
            Khan MA, Kidd BN, Lister R
  TITLE     Synthetic memory circuits for stable cell reprogramming in plants.
  JOURNAL   Nat Biotechnol. 2022 Jul 4. pii: 10.1038/s41587-022-01383-2. doi: 
            10.1038/s41587-022-01383-2.
  PUBMED    35788565
REFERENCE   2  (bases 1 to 8847)
  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: "Nat 
            Biotechnol. 2022 Jul 4. pii: 10.1038/s41587-022-01383-2. doi: 
            10.1038/s41587-022-01383-2."
COMMENT     Sequence Label: LLP727_L2_pV01_0I_TMV-rLUC_TCTP-fLUC
FEATURES             Location/Qualifiers
     source          1..8847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    824..879
                     /label=TMV Omega
                     /note="translational enhancer from the tobacco mosaic virus
                     5'-leader sequence (Gallie et al., 1988)"
     intron          911..1099
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     polyA_signal    2127..2301
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     CDS             2689..4341
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(2755..2773)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     terminator      4361..4561
                     /label=gene 7 terminator
                     /note="transcription terminator for octopine-type Ti 
                     plasmid gene 7"
     misc_feature    4625..4649
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      4737..5447
                     /label=oriV
                     /note="incP origin of replication"
     promoter        5468..5572
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5573..6367
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSLREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      6538..7126
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7027..7046
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             7462..8610
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     misc_feature    8756..8780
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 gtgccgaatt cggatccgga gtcgacaaaa tttagaacga acttaattat gatctcaaat
       61 acattgatac atatctcatc tagatctagg ttatcattat gtaagaaagt tttgacgaat
      121 atggcacgac aaaatggcta gactcgatgt aattggtatc tcaactcaac attatactta
      181 taccaaacat tagttagaca aaatttaaac aactattttt tatgtatgca agagtcagca
      241 tatgtataat tgattcagaa tcgttttgac gagttcggat gtagtagtag ccattattta
      301 atgtacatac taatcgtgaa tagtgaatat gatgaaacat tgtatcttat tgtataaata
      361 tccataaaca catcatgaaa gacactttct ttcacggtct gaattaatta tgatacaatt
      421 ctaatagaaa acgaattaaa ttacgttgaa ttgtatgaaa tctaattgaa caagccaacc
      481 acgacgacga ctaacgttgc ctggattgac tcggtttaag ttaaccacta aaaaaacgga
      541 gctgtcatgt aacacgcgga tcgagcaggt cacagtcatg aagccatcaa agcaaaagaa
      601 ctaatccaag ggctgagatg attaattagt ttaaaaatta gttaacacga gggaaaaggc
      661 tgtctgacag ccaggtcacg ttatctttac ctgtggtcga aatgattcgt gtctgtcgat
      721 tttaattatt tttttgaaag gccgaaaata aagttgtaag agataaaccc gcctatataa
      781 attcatatat tttcctctcc gctttgaata ctgtattttt acaacaatta ccaacaacaa
      841 caaacaacaa acaacattac aattactatt tacaattaca atgacttcaa aggtgtatga
      901 tcctgagcag gtaagtttct gcttctacct ttgatatata tataataatt atcattaatt
      961 agtagtaata taatatttca aatatttttt tcaaaataaa agaatgtagt atatagcaat
     1021 tgcttttctg tagtttataa gtgtgtatat tttaatttat aacttttcta atatatgacc
     1081 aaaatttgtt gatgtgcaga gaaaaagaat gataactggt ccacaatggt gggcaagatg
     1141 taaacagatg aacgtgcttg attctttcat taattactac gattcagaaa agcacgctga
     1201 gaatgcagtt atctttcttc acggaaacgc tgcatcttca tatttgtgga gacatgttgt
     1261 gcctcacatt gaaccagtgg ctaggtgtat tatcccagat cttatcggaa tgggaaagtc
     1321 tggtaaatct ggaaatggtt cttacagact tttggatcat tataagtacc tcacagcttg
     1381 gttcgaactt ctcaacttac ctaagaagat tattttcgtt ggacacgatt ggggtgcttg
     1441 cttggcattc cattactcat acgaacacca agataagatc aaagctatag ttcatgcaga
     1501 gagtgttgtg gatgtgatag aatcttggga tgagtggcca gatattgaag aggatattgc
     1561 tttgatcaag tctgaagagg gagaaaagat ggttctcgaa aataactttt tcgtggaaac
     1621 tatgcttcct tcaaagatta tgagaaaatt ggaaccagaa gagtttgctg catatctcga
     1681 acctttcaag gaaaaaggag aggttagaag gccaacactc agttggccta gggaaatacc
     1741 attagttaag ggaggtaaac ctgatgttgt gcagatcgtg agaaattata acgcttacct
     1801 tagggcatca gatgatttgc ctaagatgtt catcgagtca gatccaggat ttttcagtaa
     1861 tgctatagtt gaaggtgcaa agaaatttcc aaacaccgag ttcgttaagg tgaaaggatt
     1921 gcatttctca caagaagatg ctcctgatga gatgggaaag tatattaagt cttttgttga
     1981 aagagtgctc aaaaacgagc agggttctcg atctgatgct tctgaagtaa cgtcaacaag
     2041 ctcccagtct gaggtatgca cagtggaaac gccaggttgt gttggttctt aagcttctct
     2101 agctagagtc gatcgacaag ctcgagtttc tccataataa tgtgtgagta gttcccagat
     2161 aagggaatta gggttcctat agggtttcgc tcatgtgttg agcatataag aaacccttag
     2221 tatgtatttg tatttgtaaa atacttctat caataaaatt tctaattcct aaaaccaaaa
     2281 tccagtacta aaatccagat cgctgcaaga attcaagctt ggagccaaca ctcgaatccc
     2341 cacccgttgg accaaacccg gctcattaag cgtcggttca gatttatttc ctttatttaa
     2401 aaaaaaggaa agggtaaaaa atagaaaatt ggaaacagtt aaagcccaaa attgtaattt
     2461 accgagaatt gtaaatttac ctgaaaaccc tacgctatag tttcgactat aaataccaaa
     2521 cttaggacct cacttcagaa tcccctcgtc gctgcgtctc tctcccgcaa ccttcgattt
     2581 tcgtttattc gcatccatcg gagagagaaa acaatcaata agcgacctac tacaaacaag
     2641 taagtaagag aaaaaccaaa agaagaagag aaacaacaag aagaagtaat ggaggacgcc
     2701 aaaaacataa agaaaggccc ggcgccattc tatcctctag aggatggaac cgctggagag
     2761 caactgcata aggctatgaa gagatacgcc ctggttcctg gaacaattgc ttttacagat
     2821 gcacatatcg aggtgaacat cacgtacgcg gaatacttcg aaatgtccgt tcggttggca
     2881 gaagctatga aacgatatgg gctgaataca aatcacagaa tcgtcgtatg cagtgaaaac
     2941 tctcttcaat tctttatgcc ggtgttgggc gcgttattta tcggagttgc agttgcgccc
     3001 gcgaacgaca tttataatga acgtgaattg ctcaacagta tgaacatttc gcagcctacc
     3061 gtagtgtttg tttccaaaaa ggggttgcaa aaaattttga acgtgcaaaa aaaattacca
     3121 ataatccaga aaattattat catggattct aaaacggatt accagggatt tcagtcgatg
     3181 tacacgttcg tcacatctca tctacctccc ggttttaatg aatacgattt tgtaccagag
     3241 tcctttgatc gtgacaaaac aattgcactg ataatgaatt cctctggatc tactgggtta
     3301 cctaagggtg tggcccttcc gcatagaact gcctgcgtca gattctcgca tgccagagat
     3361 cctatttttg gcaatcaaat cattccggat actgcgattt taagtgttgt tccattccat
     3421 cacggttttg gaatgtttac tacactcgga tatttgatat gtggatttcg agtcgtctta
     3481 atgtatagat ttgaagaaga gctgttttta cgatcccttc aggattacaa aattcaaagt
     3541 gcgttgctag taccaaccct attttcattc ttcgccaaaa gcactctgat tgacaaatac
     3601 gatttatcta atttacacga aattgcttct gggggcgcac ctctttcgaa agaagtcggg
     3661 gaagcggttg caaaacgctt ccatcttcca gggatacgac aaggatatgg gctcactgag
     3721 actacatcag ctattctgat tacacccgag ggggatgata aaccgggcgc ggtcggtaaa
     3781 gttgttccat tttttgaagc gaaggttgtg gatctggata ccgggaaaac gctgggcgtt
     3841 aatcagagag gcgaattatg tgtcagagga cctatgatta tgtccggtta tgtaaacaat
     3901 ccggaagcga ccaacgcctt gattgacaag gatggatggc tacattctgg agacatagct
     3961 tactgggacg aggacgaaca cttcttcata gttgaccgct tgaagtcttt aattaaatac
     4021 aaaggatatc aggtggcccc cgctgaattg gaatcgatat tgttacaaca ccccaacatc
     4081 ttcgacgcgg gcgtggcagg tctgcccgac gatgacgccg gtgaacttcc cgccgccgtt
     4141 gttgttttgg agcacggaaa gacgatgacg gaaaaagaga tcgtggatta cgtcgccagt
     4201 caagtaacaa ccgcgaaaaa gttgcgcgga ggagttgtgt ttgtggacga agtaccgaaa
     4261 ggtcttaccg gaaaactcga cgcaagaaaa atcagagaga tcctcataaa ggccaagaag
     4321 ggcggaaagt ccaaattgta agcttgatcc cccgtcgaca gctagctata tcatcaattt
     4381 atgtattaca cataatatcg cactcagtct ttcatctacg gcaatgtacc agctgatata
     4441 atcagttatt gaaatatttc tgaatttaaa cttgcatcaa taaatttatg tttttgcttg
     4501 gactataata cctgacttgt tattttatca ataaatattt aaactatatt tctttcaaga
     4561 tcgctactag aggatgcaca tgtgaccgag ggacacgaag tgatccgttt aaactatcag
     4621 tgtttgacag gatatattgg cgggtaaacc taagagaaaa gagcgtttat tagaataatc
     4681 ggatatttaa aagggcgtga aaaggtttat ccgttcgtcc atttgtatgt gccagccgcc
     4741 tttgcgacgc tcaccgggct ggttgccctc gccgctgggc tggcggccgt ctatggccct
     4801 gcaaacgcgc cagaaacgcc gtcgaagccg tgtgcgagac accgcggccg ccggcgttgt
     4861 ggatacctcg cggaaaactt ggccctcact gacagatgag gggcggacgt tgacacttga
     4921 ggggccgact cacccggcgc ggcgttgaca gatgaggggc aggctcgatt tcggccggcg
     4981 acgtggagct ggccagcctc gcaaatcggc gaaaacgcct gattttacgc gagtttccca
     5041 cagatgatgt ggacaagcct ggggataagt gccctgcggt attgacactt gaggggcgcg
     5101 actactgaca gatgaggggc gcgatccttg acacttgagg ggcagagtgc tgacagatga
     5161 ggggcgcacc tattgacatt tgaggggctg tccacaggca gaaaatccag catttgcaag
     5221 ggtttccgcc cgtttttcgg ccaccgctaa cctgtctttt aacctgcttt taaaccaata
     5281 tttataaacc ttgtttttaa ccagggctgc gccctgtgcg cgtgaccgcg cacgccgaag
     5341 gggggtgccc ccccttctcg aaccctcccg gcccgctaac gcgggcctcc catcccccca
     5401 ggggctgcgc ccctcggccg cgaacggcct caccccaaaa atggcagcgc tggccaattc
     5461 ccgagtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     5521 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatggctaa
     5581 aatgagaata tcaccggaat tgaaaaaact gatcgaaaaa taccgctgcg taaaagatac
     5641 ggaaggaatg tctcctgcta aggtatataa gctggtggga gaaaatgaaa acctatattt
     5701 aaaaatgacg gacagccggt ataaagggac cacctatgat gtggaacggg aaaaggacat
     5761 gatgctatgg ctggaaggaa agctgcctgt tccaaaggtc ctgcactttg aacggcatga
     5821 tggctggagc aatctgctca tgagtgaggc cgatggcgtc ctttgctcgg aagagtatga
     5881 agatgaacaa agccctgaaa agattatcga gctgtatgcg gagtgcatca ggctctttca
     5941 ctccatcgac atatcggatt gtccctatac gaatagctta gacagccgct tagccgaatt
     6001 ggattactta ctgaataacg atctggccga tgtggattgc gaaaactggg aagaggacac
     6061 tccatttaaa gatccgcgcg agctgtatga ttttttaaag acggaaaagc ccgaagagga
     6121 acttgtcttt tcccacggcg acctgggaga cagcaacatc tttgtgaaag atggcaaagt
     6181 aagtggcttt attgatcttg ggagaagcgg cagggcggac aagtggtatg acattgcctt
     6241 ctgcgtccgg tcgctcaggg aggatatcgg ggaagaacag tatgtcgagc tattttttga
     6301 cttactgggg atcaagcctg attgggagaa aataaaatat tatattttac tggatgaatt
     6361 gttttagctg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     6421 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6481 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     6541 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     6601 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     6661 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     6721 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     6781 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6841 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6901 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     6961 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     7021 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     7081 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     7141 ggccttttta cggttcctgg cagatcctag atgtggcgca acgatgccgg cgacaagcag
     7201 gagcgcaccg acttcttccg catcaagtgt tttggctctc aggccgaggc ccacggcaag
     7261 tatttgggca aggggtcgct ggtattcgtg cagggcaaga ttcggaatac caagtacgag
     7321 aaggacggcc agacggtcta cgggaccgac ttcattgccg ataaggtgga ttatctggac
     7381 accaaggcac caggcgggtc aaatcaggaa taagggcaca ttgccccggc gtgagtcggg
     7441 gcaatcccgc aaggagggtg aatgaatcgg acgtttgacc ggaaggcata caggcaagaa
     7501 ctgatcgacg cggggttttc cgccgaggat gccgaaacca tcgcaagccg caccgtcatg
     7561 cgtgcgcccc gcgaaacctt ccagtccgtc ggctcgatgg tccagcaagc tacggccaag
     7621 atcgagcgcg acagcgtgca actggctccc cctgccctgc ccgcgccatc ggccgccgtg
     7681 gagcgttcgc gtcgtcttga acaggaggcg gcaggtttgg cgaagtcgat gaccatcgac
     7741 acgcgaggaa ctatgacgac caagaagcga aaaaccgccg gcgaggacct ggcaaaacag
     7801 gtcagcgagg ccaagcaggc cgcgttgctg aaacacacga agcagcagat caaggaaatg
     7861 cagctttcct tgttcgatat tgcgccgtgg ccggacacga tgcgagcgat gccaaacgac
     7921 acggcccgct ctgccctgtt caccacgcgc aacaagaaaa tcccgcgcga ggcgctgcaa
     7981 aacaaggtca ttttccacgt caacaaggac gtgaagatca cctacaccgg cgtcgagctg
     8041 cgggccgacg atgacgaact ggtgtggcag caggtgttgg agtacgcgaa gcgcacccct
     8101 atcggcgagc cgatcacctt cacgttctac gagctttgcc aggacctggg ctggtcgatc
     8161 aatggccggt attacacgaa ggccgaggaa tgcctgtcgc gcctacaggc gacggcgatg
     8221 ggcttcacgt ccgaccgcgt tgggcacctg gaatcggtgt cgctgctgca ccgcttccgc
     8281 gtcctggacc gtggcaagaa aacgtcccgt tgccaggtcc tgatcgacga ggaaatcgtc
     8341 gtgctgtttg ctggcgacca ctacacgaaa ttcatatggg agaagtaccg caagctgtcg
     8401 ccgacggccc gacggatgtt cgactatttc agctcgcacc gggagccgta cccgctcaag
     8461 ctggaaacct tccgcctcat gtgcggatcg gattccaccc gcgtgaagaa gtggcgcgag
     8521 caggtcggcg aagcctgcga agagttgcga ggcagcggcc tggtggaaca cgcctgggtc
     8581 aatgatgacc tggtgcattg caaacgctag ggccttgtgg ggtcagttcc ggctgggggt
     8641 tcagcagccc ctgctcggat ctgttggacc ggacagtagt catggttgat gggctgcctg
     8701 tatcgagtgg tgattttgtg ccgagctgcc ggtcggggag ctgttggctg gctggtggca
     8761 ggatatattg tggtgtaaac aaattgacgc ttagacaact taataacaca ttgcggacgt
     8821 ttttaatgta ctggggttga acactct
//
