LOCUS       sequence-490733-        7279 bp    DNA     circular SYN 27-OCT-2025
DEFINITION  TCTP-SynPro-12 engineered promoter driving the expression of Renilla
            luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7279)
  AUTHORS   Khan MA, Herring G, Zhu JY, Oliva M, Fourie E, Johnston B, Zhang Z, 
            Potter J, Pineda L, Pflueger J, Swain T, Pflueger C, Lloyd JPB, 
            Secco D, Small I, Kidd BN, Lister R
  TITLE     CRISPRi-based circuits to control gene expression in plants.
  JOURNAL   Nat Biotechnol. 2025 Mar;43(3):416-430. doi: 
            10.1038/s41587-024-02236-w. Epub 2024 May 20.
  PUBMED    38769424
REFERENCE   2  (bases 1 to 7279)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 27, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41587-024-02236-w"; journalName: "Nat Biotechnol"; date: 
            "2025-03"; volume: "43"; issue: "3"; pages: "416-430"
COMMENT     Sequence Label: LLP959
FEATURES             Location/Qualifiers
     source          1..7279
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          1589..1777
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     primer_bind     1777..1795
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     promoter        complement(1872..2051)
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     misc_feature    2065..2156
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     polyA_signal    2170..2218
                     /note="synthetic polyadenylation signal"
     intron          3450..3638
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     primer_bind     complement(4037..4056)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4038..4056)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4063..4080)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4063..4079)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4072..4094)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4221..4676
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4308..4327)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4518..4539
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4702..4806
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4807..4875,4876..5667)
                     /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     complement(5025..5044)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5838..6426
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6327..6346
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6580..6597
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6714..6735
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6750..6755,6756..6773,6774..6780)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6788..6804
                     /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     6793..6815
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6812..6828
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6812..6828
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6849..6867
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    6886..6977
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     polyA_signal    6991..7039
                     /note="synthetic polyadenylation signal"
     terminator      complement(join(7040..7279,1..13))
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
ORIGIN
        1 tgtttgaacg atcctcgagt tacacggcga tctttccgcc cttcttggcc tttatgagga
       61 tctctctgat ttttcttgcg tcgagttttc cggtaagacc tttcggtact tcgtccacaa
      121 acacaactcc tccgcgcaac tttttcgcgg ttgttacttg actggcgacg taatccacga
      181 tctctttttc cgtcatcgtc tttccgtgct ccaaaacaac aacggcggcg ggaagttcac
      241 cggcgtcatc gtcgggaaga cctgcgacac ctgcgtcgaa gatgttgggg tgttggagca
      301 agatggattc caattcagcg ggagccacct gatagccttt gtacttaatc agagacttca
      361 ggcggtcaac gatgaagaag tgttcgtctt cgtcccagta agctatgtct ccagaatgta
      421 gccatccatc cttgtcaatc aaggcgttgg tcgcttccgg attgtttaca taaccggaca
      481 taatcatagg acctctcaca cacagttcgc ctctttgatt aacgcccagc gttttcccgg
      541 tatccagatc cacaaccttc gcttcaaaaa atggaacaac tttaccgacc gcgcccggtt
      601 tatcatcccc ctcgggtgta atcagaatag ctgatgtagt ctcagtgagc ccatatcctt
      661 gcctgatacc tggcagatgg aacctcttgg caaccgcttc cccgacttcc ttagagaggg
      721 gagcgccacc agaagcaatt tcgtgtaaat tagataaatc gtatttgtca atcagagtgc
      781 ttttggcgaa gaaggagaat agggttggca ccagcagcgc actttgaatc ttgtaatcct
      841 gaaggctcct cagaaacagc tcttcttcaa atctatacat taagacgact cgaaatccac
      901 atatcaaata tccgagtgta gtaaacattc caaaaccgtg atggaatgga acaacactta
      961 aaatcgcagt atccggaatg atttgattgc caaaaatagg atctctggca tgcgagaatc
     1021 tcacgcaggc agttctatga ggcagagcga cacctttagg cagaccagta gatccagagg
     1081 agttcatgat cagtgcaatt gtcttgtccc tatcgaagga ctctggcaca aaatcgtatt
     1141 cattaaaacc gggaggtaga tgagatgtga cgaacgtgta catcgactga aatccctggt
     1201 aatccgtttt agaatccatg ataataattt tttggatgat tgggagcttt ttttgcacgt
     1261 tcaaaatttt ttgcaacccc tttttggaaa cgaacaccac ggtaggctgc gaaatgccca
     1321 tactgttgag caattcacgt tcattataaa tgtcgttcgc gggcgcaact gcaactccga
     1381 taaataacgc gcccaacacc ggcataaaga attgaagaga gttttcactg catacgacga
     1441 ttctgtgatt tgtattcagc ccatatcgtt tcatagcttc tgccaaccga acggacattt
     1501 cgaagtactc agcgtaagtg atgtccacct cgatatgtgc atctgtaaaa gcaattgttc
     1561 caggaaccag ggcgtatctc ttcatagcct gcacatcaac aaattttggt catatattag
     1621 aaaagttata aattaaaata tacacactta taaactacag aaaagcaatt gctatatact
     1681 acattctttt attttgaaaa aaatatttga aatattatat tactactaat taatgataat
     1741 tattatatat atatcaaagg tagaagcaga aacttacctt atgcagttgc tctccagcgg
     1801 ttccatcttc cagcggatag aatggcgccg ggcctttctt tatgtttttg gcgtcttcca
     1861 tccatggact ctaattggat accgagggga atttatggaa cgtcagtgga gcatttttga
     1921 caagaaatat ttgctagctg atagtgacct taggcgactt ttgaacgcgc aataatggtt
     1981 tctgacgtat gtgcttagct cattaaactc cagaaacccg cggctcagtg gctccttcaa
     2041 cgttgcggtt ctctagagag cgctagagaa atgttctggc acctgcactt gcactgggga
     2101 cagcctattt tgctagtttg ttttgtttcg ttttgttttg atggagagcg tatgttagta
     2161 ctatcgattc acacaaaaaa ccaacacaca gatgtaatga aaataaagat attttattac
     2221 tagtaggtca ctggattttg gttttaggaa ttagaaattt tattgataga agtattttac
     2281 aaatacaaat acatactaag ggtttcttat atgctcaaca catgagcgaa accctataag
     2341 aaccctaatt cccttatctg ggaactactc acacattatt ctggagaaaa atagagagag
     2401 atagatttgt agagagagac tggtgatttt tgcggactct agcatggccg ggtaccttaa
     2461 gaaccaacac aacctggcgt ttccactgtg catacctcag actgggagct tgttgacgtt
     2521 acttcagaag catcagatcg agaaccctgc tcgtttttga gcactctttc aacaaaagac
     2581 ttaatatact ttcccatctc atcaggagca tcttcttgtg agaaatgcaa tcctttcacc
     2641 ttaacgaact cggtgtttgg aaatttcttt gcaccttcaa ctatagcatt actgaaaaat
     2701 cctggatctg actcgatgaa catcttaggc aaatcatctg atgccctaag gtaagcgtta
     2761 taatttctca cgatctgcac aacatcaggt ttacctccct taactaatgg tatttcccta
     2821 ggccaactga gtgttggcct tctaacctct cctttttcct tgaaaggttc gagatatgca
     2881 gcaaactctt ctggttccaa ttttctcata atctttgaag gaagcatagt ttccacgaaa
     2941 aagttatttt cgagaaccat cttttctccc tcttcagact tgatcaaagc aatatcctct
     3001 tcaatatctg gccactcatc ccaagattct atcacatcca caacactctc tgcatgaact
     3061 atagctttga tcttatcttg gtgttcgtat gagtaatgga atgccaagca agcaccccaa
     3121 tcgtgtccaa cgaaaataat cttcttaggt aagttgagaa gttcgaacca agctgtgagg
     3181 tacttataat gatccaaaag tctgtaagaa ccatttccag atttaccaga ctttcccatt
     3241 ccgataagat ctgggataat acacctagcc actggttcaa tgtgaggcac aacatgtctc
     3301 cacaaatatg aagatgcagc gtttccgtga agaaagataa ctgcattctc agcgtgcttt
     3361 tctgaatcgt agtaattaat gaaagaatca agcacgttca tctgtttaca tcttgcccac
     3421 cattgtggac cagttatcat tctttttctc tgcacatcaa caaattttgg tcatatatta
     3481 gaaaagttat aaattaaaat atacacactt ataaactaca gaaaagcaat tgctatatac
     3541 tacattcttt tattttgaaa aaaatatttg aaatattata ttactactaa ttaatgataa
     3601 ttattatata tatatcaaag gtagaagcag aaacttacct gctcaggatc atacaccttt
     3661 gaagtcatct gcagttatgt cgacggatcc ggtcgcttat tgattgtttt ctctctccga
     3721 tggatgcgaa taaacgaaaa tcgaaggttg cgggagagag acgcagcgac gaggggattc
     3781 cactctaaac ctatacgtaa agtatttata gtccagacat ggagcaagtt accacaggta
     3841 aatttacaat tctcggtaaa ttacaatttt gggctttaac tgtttccaat tttctatttt
     3901 ttaccctttc ctttttttta aataaaggaa ataaatctga accgacgctt aatgagccgg
     3961 gtttggtcca acgggtgggg attcgagtgt tggaacgcgt tcggaattcg gcatgatcaa
     4021 gcttgtaccc aattcgccct atagtgagtc gtattacaat tcactggccg tcgttttaca
     4081 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     4141 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     4201 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     4261 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     4321 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     4381 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     4441 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     4501 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     4561 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     4621 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt
     4681 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     4741 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4801 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4861 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     4921 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     4981 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5041 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5101 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5161 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5221 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5281 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5341 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     5401 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     5461 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     5521 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     5581 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     5641 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     5701 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     5761 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     5821 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5881 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5941 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6001 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6061 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6121 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6181 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6241 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6301 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6361 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6421 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6481 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     6541 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     6601 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     6661 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     6721 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     6781 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     6841 aagctcgaaa ttaaccctca ctaaagggaa caaaagctgg agctcagaga aatgttctgg
     6901 cacctgcact tgcactgggg acagcctatt ttgctagttt gttttgtttc gttttgtttt
     6961 gatggagagc gtatgttagt actatcgatt cacacaaaaa accaacacac agatgtaatg
     7021 aaaataaaga tattttattg atctagtaac atagatgaca ccgcgcgcga taatttatcc
     7081 tagtttgcgc gctatatttt gttttctatc gcgtattaaa tgtataattg cgggactcta
     7141 atcataaaaa cccatctcat aaataacgtc atgcattaca tgttaattat tacatgctta
     7201 acgtaattca acagaaatta tatgataatc gtcgcaagac cggcaacagg attcaatctt
     7261 aagaaacttt attgccaaa
//
