LOCUS       sequence-490460-        7320 bp    DNA     circular SYN 23-OCT-2025
DEFINITION  CaMV 35S-SynPro-07 engineered promoter driving the expression of 
            Renilla luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7320)
  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 7320)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 23, 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: LLP972
FEATURES             Location/Qualifiers
     source          1..7320
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          1085..1273
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     polyA_signal    2505..2553
                     /note="synthetic polyadenylation signal"
     misc_feature    2567..2658
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     promoter        2672..2851
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     primer_bind     complement(2928..2946)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     intron          2946..3134
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     terminator      4710..4962
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     polyA_signal    4963..5011
                     /note="synthetic polyadenylation signal"
     misc_feature    5025..5116
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     promoter        complement(5135..5153)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5174..5190)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5174..5190)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5187..5209)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5198..5214
                     /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(5222..5228,5229..5246,5247..5252))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5267..5288
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5405..5422)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5576..6164)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5656..5675)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6335..7126,7127..7195))
                     /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     6958..6977
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7196..7300)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtacaag
      661 cttgatcatg ccgaattccg aacgcgttga gacttttcaa caaagggtaa tatcgggaaa
      721 cctcctcgga ttccattgcc cagctatctg tcacttcatc aaaaggacag tagaaaagga
      781 aggtggcacc tacaaatgcc atcattgcga taaaggaaag gctatcgttc aagatgcctc
      841 tgccgacagt ggtcccaaag atggaccccc acccacgagg agcatcgtgg aaaaagaaga
      901 cgttccaacc acgtcttcaa agcaagtgga ttgatgtgat atctccactg acgtaaggga
      961 tgacgcacaa tcccactggg taggagagac tcacgctggt atataaggag gcagtcccgg
     1021 ctcgctgggg acggatccgt cgacataact gcagatgact tcaaaggtgt atgatcctga
     1081 gcaggtaagt ttctgcttct acctttgata tatatataat aattatcatt aattagtagt
     1141 aatataatat ttcaaatatt tttttcaaaa taaaagaatg tagtatatag caattgcttt
     1201 tctgtagttt ataagtgtgt atattttaat ttataacttt tctaatatat gaccaaaatt
     1261 tgttgatgtg cagagaaaaa gaatgataac tggtccacaa tggtgggcaa gatgtaaaca
     1321 gatgaacgtg cttgattctt tcattaatta ctacgattca gaaaagcacg ctgagaatgc
     1381 agttatcttt cttcacggaa acgctgcatc ttcatatttg tggagacatg ttgtgcctca
     1441 cattgaacca gtggctaggt gtattatccc agatcttatc ggaatgggaa agtctggtaa
     1501 atctggaaat ggttcttaca gacttttgga tcattataag tacctcacag cttggttcga
     1561 acttctcaac ttacctaaga agattatttt cgttggacac gattggggtg cttgcttggc
     1621 attccattac tcatacgaac accaagataa gatcaaagct atagttcatg cagagagtgt
     1681 tgtggatgtg atagaatctt gggatgagtg gccagatatt gaagaggata ttgctttgat
     1741 caagtctgaa gagggagaaa agatggttct cgaaaataac tttttcgtgg aaactatgct
     1801 tccttcaaag attatgagaa aattggaacc agaagagttt gctgcatatc tcgaaccttt
     1861 caaggaaaaa ggagaggtta gaaggccaac actcagttgg cctagggaaa taccattagt
     1921 taagggaggt aaacctgatg ttgtgcagat cgtgagaaat tataacgctt accttagggc
     1981 atcagatgat ttgcctaaga tgttcatcga gtcagatcca ggatttttca gtaatgctat
     2041 agttgaaggt gcaaagaaat ttccaaacac cgagttcgtt aaggtgaaag gattgcattt
     2101 ctcacaagaa gatgctcctg atgagatggg aaagtatatt aagtcttttg ttgaaagagt
     2161 gctcaaaaac gagcagggtt ctcgatctga tgcttctgaa gtaacgtcaa caagctccca
     2221 gtctgaggta tgcacagtgg aaacgccagg ttgtgttggt tcttaaggta cccggccatg
     2281 ctagagtccg caaaaatcac cagtctctct ctacaaatct atctctctct atttttctcc
     2341 agaataatgt gtgagtagtt cccagataag ggaattaggg ttcttatagg gtttcgctca
     2401 tgtgttgagc atataagaaa cccttagtat gtatttgtat ttgtaaaata cttctatcaa
     2461 taaaatttct aattcctaaa accaaaatcc agtgacctac tagtaataaa atatctttat
     2521 tttcattaca tctgtgtgtt ggttttttgt gtgaatcgat agtactaaca tacgctctcc
     2581 atcaaaacaa aacgaaacaa aacaaactag caaaataggc tgtccccagt gcaagtgcag
     2641 gtgccagaac atttctctag cgctctctag agaaccgcaa cgttgaagga gccactgagc
     2701 cgcgggtttc tggagtttaa tgagctaagc acatacgtca gaaaccatta ttgcgcgttc
     2761 aaaagtcgcc taaggtcact atcagctagc aaatatttct tgtcaaaaat gctccactga
     2821 cgttccataa attcccctcg gtatccaatt agagtccatg gatggaagac gccaaaaaca
     2881 taaagaaagg cccggcgcca ttctatccgc tggaagatgg aaccgctgga gagcaactgc
     2941 ataaggtaag tttctgcttc tacctttgat atatatataa taattatcat taattagtag
     3001 taatataata tttcaaatat ttttttcaaa ataaaagaat gtagtatata gcaattgctt
     3061 ttctgtagtt tataagtgtg tatattttaa tttataactt ttctaatata tgaccaaaat
     3121 ttgttgatgt gcaggctatg aagagatacg ccctggttcc tggaacaatt gcttttacag
     3181 atgcacatat cgaggtggac atcacttacg ctgagtactt cgaaatgtcc gttcggttgg
     3241 cagaagctat gaaacgatat gggctgaata caaatcacag aatcgtcgta tgcagtgaaa
     3301 actctcttca attctttatg ccggtgttgg gcgcgttatt tatcggagtt gcagttgcgc
     3361 ccgcgaacga catttataat gaacgtgaat tgctcaacag tatgggcatt tcgcagccta
     3421 ccgtggtgtt cgtttccaaa aaggggttgc aaaaaatttt gaacgtgcaa aaaaagctcc
     3481 caatcatcca aaaaattatt atcatggatt ctaaaacgga ttaccaggga tttcagtcga
     3541 tgtacacgtt cgtcacatct catctacctc ccggttttaa tgaatacgat tttgtgccag
     3601 agtccttcga tagggacaag acaattgcac tgatcatgaa ctcctctgga tctactggtc
     3661 tgcctaaagg tgtcgctctg cctcatagaa ctgcctgcgt gagattctcg catgccagag
     3721 atcctatttt tggcaatcaa atcattccgg atactgcgat tttaagtgtt gttccattcc
     3781 atcacggttt tggaatgttt actacactcg gatatttgat atgtggattt cgagtcgtct
     3841 taatgtatag atttgaagaa gagctgtttc tgaggagcct tcaggattac aagattcaaa
     3901 gtgcgctgct ggtgccaacc ctattctcct tcttcgccaa aagcactctg attgacaaat
     3961 acgatttatc taatttacac gaaattgctt ctggtggcgc tcccctctct aaggaagtcg
     4021 gggaagcggt tgccaagagg ttccatctgc caggtatcag gcaaggatat gggctcactg
     4081 agactacatc agctattctg attacacccg agggggatga taaaccgggc gcggtcggta
     4141 aagttgttcc attttttgaa gcgaaggttg tggatctgga taccgggaaa acgctgggcg
     4201 ttaatcaaag aggcgaactg tgtgtgagag gtcctatgat tatgtccggt tatgtaaaca
     4261 atccggaagc gaccaacgcc ttgattgaca aggatggatg gctacattct ggagacatag
     4321 cttactggga cgaagacgaa cacttcttca tcgttgaccg cctgaagtct ctgattaagt
     4381 acaaaggcta tcaggtggct cccgctgaat tggaatccat cttgctccaa caccccaaca
     4441 tcttcgacgc aggtgtcgca ggtcttcccg acgatgacgc cggtgaactt cccgccgccg
     4501 ttgttgtttt ggagcacgga aagacgatga cggaaaaaga gatcgtggat tacgtcgcca
     4561 gtcaagtaac aaccgcgaaa aagttgcgcg gaggagttgt gtttgtggac gaagtaccga
     4621 aaggtcttac cggaaaactc gacgcaagaa aaatcagaga gatcctcata aaggccaaga
     4681 agggcggaaa gatcgccgtg taactcgagg atcgttcaaa catttggcaa taaagtttct
     4741 taagattgaa tcctgttgcc ggtcttgcga cgattatcat ataatttctg ttgaattacg
     4801 ttaagcatgt aataattaac atgtaatgca tgacgttatt tatgagatgg gtttttatga
     4861 ttagagtccc gcaattatac atttaatacg cgatagaaaa caaaatatag cgcgcaaact
     4921 aggataaatt atcgcgcgcg gtgtcatcta tgttactaga tcaataaaat atctttattt
     4981 tcattacatc tgtgtgttgg ttttttgtgt gaatcgatag tactaacata cgctctccat
     5041 caaaacaaaa cgaaacaaaa caaactagca aaataggctg tccccagtgc aagtgcaggt
     5101 gccagaacat ttctctgagc tccagctttt gttcccttta gtgagggtta atttcgagct
     5161 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5221 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5281 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5341 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     5401 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5461 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5521 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5581 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5641 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5701 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5761 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5821 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5881 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5941 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6001 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6061 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6121 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6181 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6241 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6301 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6361 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6421 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6481 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6541 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6601 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6661 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6721 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6781 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6841 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6901 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6961 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     7021 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7081 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7141 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7201 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7261 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
//
