LOCUS       sequence-490745-        7486 bp    DNA     circular SYN 27-OCT-2025
DEFINITION  CaMV 35S-SynPro-01 engineered promoter driving the expression of 
            Renilla luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7486)
  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 7486)
  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: LLP1024
FEATURES             Location/Qualifiers
     source          1..7486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    967..1015
                     /note="synthetic polyadenylation signal"
     misc_feature    1029..1120
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     promoter        1134..1478
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1422..1442
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     primer_bind     complement(1555..1573)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     intron          1573..1761
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     terminator      3337..3589
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     polyA_signal    3590..3638
                     /note="synthetic polyadenylation signal"
     misc_feature    3652..3743
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     promoter        complement(3762..3780)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3801..3817)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3801..3817)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3814..3836)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3825..3841
                     /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)."
     promoter        complement(join(3849..3855,3856..3873,3874..3879))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3894..3915
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4032..4049)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4203..4791)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4283..4302)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4962..5753,5754..5822))
                     /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     5585..5604
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5823..5927)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5953..6408)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6090..6111)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6302..6321
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6535..6557
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6549..6566
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6550..6566
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6573..6592
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6573..6591
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          7033..7221
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
ORIGIN
        1 agtacctcac agcttggttc gaacttctca acttacctaa gaagattatt ttcgttggac
       61 acgattgggg tgcttgcttg gcattccatt actcatacga acaccaagat aagatcaaag
      121 ctatagttca tgcagagagt gttgtggatg tgatagaatc ttgggatgag tggccagata
      181 ttgaagagga tattgctttg atcaagtctg aagagggaga aaagatggtt ctcgaaaata
      241 actttttcgt ggaaactatg cttccttcaa agattatgag aaaattggaa ccagaagagt
      301 ttgctgcata tctcgaacct ttcaaggaaa aaggagaggt tagaaggcca acactcagtt
      361 ggcctaggga aataccatta gttaagggag gtaaacctga tgttgtgcag atcgtgagaa
      421 attataacgc ttaccttagg gcatcagatg atttgcctaa gatgttcatc gagtcagatc
      481 caggattttt cagtaatgct atagttgaag gtgcaaagaa atttccaaac accgagttcg
      541 ttaaggtgaa aggattgcat ttctcacaag aagatgctcc tgatgagatg ggaaagtata
      601 ttaagtcttt tgttgaaaga gtgctcaaaa acgagcaggg ttctcgatct gatgcttctg
      661 aagtaacgtc aacaagctcc cagtctgagg tatgcacagt ggaaacgcca ggttgtgttg
      721 gttcttaagg tacccggcca tgctagagtc cgcaaaaatc accagtctct ctctacaaat
      781 ctatctctct ctatttttct ccagaataat gtgtgagtag ttcccagata agggaattag
      841 ggttcttata gggtttcgct catgtgttga gcatataaga aacccttagt atgtatttgt
      901 atttgtaaaa tacttctatc aataaaattt ctaattccta aaaccaaaat ccagtgacct
      961 actagtaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg
     1021 atagtactaa catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag
     1081 gctgtcccca gtgcaagtgc aggtgccaga acatttctct agcgctctct agatgagact
     1141 tttcaacaaa gggtaatatc gggaaacctc ctcggattcc attgcccagc tatctgtcac
     1201 ttcatcaaaa ggacagtaga aaaggaaggt ggcacctaca aatgccatca ttgcgataaa
     1261 ggaaaggcta tcgttcaaga tgcctctgcc gacagtggtc ccaaagatgg acccccaccc
     1321 acgaggagca tcgtggaaaa agaagacgtt ccaaccacgt cttcaaagca agtggattga
     1381 tgtgatatct ccactgacgt aagggatgac gcacaatccc actatccttc gcaagaccct
     1441 tcctctatat aaggaagttc atttcatttg gagaggacga gtccatggat ggaagacgcc
     1501 aaaaacataa agaaaggccc ggcgccattc tatccgctgg aagatggaac cgctggagag
     1561 caactgcata aggtaagttt ctgcttctac ctttgatata tatataataa ttatcattaa
     1621 ttagtagtaa tataatattt caaatatttt tttcaaaata aaagaatgta gtatatagca
     1681 attgcttttc tgtagtttat aagtgtgtat attttaattt ataacttttc taatatatga
     1741 ccaaaatttg ttgatgtgca ggctatgaag agatacgccc tggttcctgg aacaattgct
     1801 tttacagatg cacatatcga ggtggacatc acttacgctg agtacttcga aatgtccgtt
     1861 cggttggcag aagctatgaa acgatatggg ctgaatacaa atcacagaat cgtcgtatgc
     1921 agtgaaaact ctcttcaatt ctttatgccg gtgttgggcg cgttatttat cggagttgca
     1981 gttgcgcccg cgaacgacat ttataatgaa cgtgaattgc tcaacagtat gggcatttcg
     2041 cagcctaccg tggtgttcgt ttccaaaaag gggttgcaaa aaattttgaa cgtgcaaaaa
     2101 aagctcccaa tcatccaaaa aattattatc atggattcta aaacggatta ccagggattt
     2161 cagtcgatgt acacgttcgt cacatctcat ctacctcccg gttttaatga atacgatttt
     2221 gtgccagagt ccttcgatag ggacaagaca attgcactga tcatgaactc ctctggatct
     2281 actggtctgc ctaaaggtgt cgctctgcct catagaactg cctgcgtgag attctcgcat
     2341 gccagagatc ctatttttgg caatcaaatc attccggata ctgcgatttt aagtgttgtt
     2401 ccattccatc acggttttgg aatgtttact acactcggat atttgatatg tggatttcga
     2461 gtcgtcttaa tgtatagatt tgaagaagag ctgtttctga ggagccttca ggattacaag
     2521 attcaaagtg cgctgctggt gccaacccta ttctccttct tcgccaaaag cactctgatt
     2581 gacaaatacg atttatctaa tttacacgaa attgcttctg gtggcgctcc cctctctaag
     2641 gaagtcgggg aagcggttgc caagaggttc catctgccag gtatcaggca aggatatggg
     2701 ctcactgaga ctacatcagc tattctgatt acacccgagg gggatgataa accgggcgcg
     2761 gtcggtaaag ttgttccatt ttttgaagcg aaggttgtgg atctggatac cgggaaaacg
     2821 ctgggcgtta atcaaagagg cgaactgtgt gtgagaggtc ctatgattat gtccggttat
     2881 gtaaacaatc cggaagcgac caacgccttg attgacaagg atggatggct acattctgga
     2941 gacatagctt actgggacga agacgaacac ttcttcatcg ttgaccgcct gaagtctctg
     3001 attaagtaca aaggctatca ggtggctccc gctgaattgg aatccatctt gctccaacac
     3061 cccaacatct tcgacgcagg tgtcgcaggt cttcccgacg atgacgccgg tgaacttccc
     3121 gccgccgttg ttgttttgga gcacggaaag acgatgacgg aaaaagagat cgtggattac
     3181 gtcgccagtc aagtaacaac cgcgaaaaag ttgcgcggag gagttgtgtt tgtggacgaa
     3241 gtaccgaaag gtcttaccgg aaaactcgac gcaagaaaaa tcagagagat cctcataaag
     3301 gccaagaagg gcggaaagat cgccgtgtaa ctcgaggatc gttcaaacat ttggcaataa
     3361 agtttcttaa gattgaatcc tgttgccggt cttgcgacga ttatcatata atttctgttg
     3421 aattacgtta agcatgtaat aattaacatg taatgcatga cgttatttat gagatgggtt
     3481 tttatgatta gagtcccgca attatacatt taatacgcga tagaaaacaa aatatagcgc
     3541 gcaaactagg ataaattatc gcgcgcggtg tcatctatgt tactagatca ataaaatatc
     3601 tttattttca ttacatctgt gtgttggttt tttgtgtgaa tcgatagtac taacatacgc
     3661 tctccatcaa aacaaaacga aacaaaacaa actagcaaaa taggctgtcc ccagtgcaag
     3721 tgcaggtgcc agaacatttc tctgagctcc agcttttgtt ccctttagtg agggttaatt
     3781 tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     3841 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     3901 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     3961 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4021 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4081 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4141 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4201 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4261 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4321 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4381 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4441 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4501 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4561 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4621 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     4681 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4741 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4801 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     4861 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     4921 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4981 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5041 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5101 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5161 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5221 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5281 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5341 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5401 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5461 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5521 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5581 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5641 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5701 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5761 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5821 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5881 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     5941 aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt
     6001 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     6061 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     6121 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     6181 aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     6241 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     6301 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     6361 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat
     6421 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
     6481 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     6541 cacgacgttg taaaacgacg gccagtgaat tgtaatacga ctcactatag ggcgaattgg
     6601 gtacaagctt gatcatgccg aattccgaac gcgtttgaga cttttcaaca aagggtaata
     6661 tcgggaaacc tcctcggatt ccattgccca gctatctgtc acttcatcaa aaggacagta
     6721 gaaaaggaag gtggcaccta caaatgccat cattgcgata aaggaaaggc tatcgttcaa
     6781 gatgcctctg ccgacagtgg tcccaaagat ggacccccac ccacgaggag catcgtggaa
     6841 aaagaagacg ttccaaccac gtcttcaaag caagtggatt gatgtgatat ctccactgac
     6901 gtaagggatg acgcacaatc ccactataga accgatcctc ccattggtat ataatacctc
     6961 atcaggaaca tgttggggac ggatccgtcg acataactgc agatgacttc aaaggtgtat
     7021 gatcctgagc aggtaagttt ctgcttctac ctttgatata tatataataa ttatcattaa
     7081 ttagtagtaa tataatattt caaatatttt tttcaaaata aaagaatgta gtatatagca
     7141 attgcttttc tgtagtttat aagtgtgtat attttaattt ataacttttc taatatatga
     7201 ccaaaatttg ttgatgtgca gagaaaaaga atgataactg gtccacaatg gtgggcaaga
     7261 tgtaaacaga tgaacgtgct tgattctttc attaattact acgattcaga aaagcacgct
     7321 gagaatgcag ttatctttct tcacggaaac gctgcatctt catatttgtg gagacatgtt
     7381 gtgcctcaca ttgaaccagt ggctaggtgt attatcccag atcttatcgg aatgggaaag
     7441 tctggtaaat ctggaaatgg ttcttacaga cttttggatc attata
//
