LOCUS       sequence-490730-        7279 bp    DNA     circular SYN 27-OCT-2025
DEFINITION  TCTP-SynPro-03 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: LLP950
FEATURES             Location/Qualifiers
     source          1..7279
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1029..1077
                     /note="synthetic polyadenylation signal"
     misc_feature    1091..1182
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     promoter        1196..1375
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     primer_bind     complement(1452..1470)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     intron          1470..1658
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     terminator      3234..3486
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     polyA_signal    3487..3535
                     /note="synthetic polyadenylation signal"
     misc_feature    3549..3640
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     promoter        complement(3659..3677)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3698..3714)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3698..3714)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3711..3733)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3722..3738
                     /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(3746..3752,3753..3770,3771..3776))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3791..3812
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3929..3946)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4100..4688)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4180..4199)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4859..5650,5651..5719))
                     /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     5482..5501
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5720..5824)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5850..6305)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5987..6008)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6199..6218
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6432..6454
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6446..6463
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6447..6463
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6470..6489
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6470..6488
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          6888..7076
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
ORIGIN
        1 tatcggaatg ggaaagtctg gtaaatctgg aaatggttct tacagacttt tggatcatta
       61 taagtacctc acagcttggt tcgaacttct caacttacct aagaagatta ttttcgttgg
      121 acacgattgg ggtgcttgct tggcattcca ttactcatac gaacaccaag ataagatcaa
      181 agctatagtt catgcagaga gtgttgtgga tgtgatagaa tcttgggatg agtggccaga
      241 tattgaagag gatattgctt tgatcaagtc tgaagaggga gaaaagatgg ttctcgaaaa
      301 taactttttc gtggaaacta tgcttccttc aaagattatg agaaaattgg aaccagaaga
      361 gtttgctgca tatctcgaac ctttcaagga aaaaggagag gttagaaggc caacactcag
      421 ttggcctagg gaaataccat tagttaaggg aggtaaacct gatgttgtgc agatcgtgag
      481 aaattataac gcttacctta gggcatcaga tgatttgcct aagatgttca tcgagtcaga
      541 tccaggattt ttcagtaatg ctatagttga aggtgcaaag aaatttccaa acaccgagtt
      601 cgttaaggtg aaaggattgc atttctcaca agaagatgct cctgatgaga tgggaaagta
      661 tattaagtct tttgttgaaa gagtgctcaa aaacgagcag ggttctcgat ctgatgcttc
      721 tgaagtaacg tcaacaagct cccagtctga ggtatgcaca gtggaaacgc caggttgtgt
      781 tggttcttaa ggtacccggc catgctagag tccgcaaaaa tcaccagtct ctctctacaa
      841 atctatctct ctctattttt ctccagaata atgtgtgagt agttcccaga taagggaatt
      901 agggttctta tagggtttcg ctcatgtgtt gagcatataa gaaaccctta gtatgtattt
      961 gtatttgtaa aatacttcta tcaataaaat ttctaattcc taaaaccaaa atccagtgac
     1021 ctactagtaa taaaatatct ttattttcat tacatctgtg tgttggtttt ttgtgtgaat
     1081 cgatagtact aacatacgct ctccatcaaa acaaaacgaa acaaaacaaa ctagcaaaat
     1141 aggctgtccc cagtgcaagt gcaggtgcca gaacatttct ctagcgctct ctagagaacc
     1201 gcaacgttga aggagccact gagccgcggg tttctggagt ttaatgagct aagcacatac
     1261 gtcagaaacc attattgcgc gttcaaaagt cgcctaaggt cactatcagc tagcaaatat
     1321 ttcttgtcaa aaatgctcca ctgacgttcc ataaattccc ctcggtatcc aattagagtc
     1381 catggatgga agacgccaaa aacataaaga aaggcccggc gccattctat ccgctggaag
     1441 atggaaccgc tggagagcaa ctgcataagg taagtttctg cttctacctt tgatatatat
     1501 ataataatta tcattaatta gtagtaatat aatatttcaa atattttttt caaaataaaa
     1561 gaatgtagta tatagcaatt gcttttctgt agtttataag tgtgtatatt ttaatttata
     1621 acttttctaa tatatgacca aaatttgttg atgtgcaggc tatgaagaga tacgccctgg
     1681 ttcctggaac aattgctttt acagatgcac atatcgaggt ggacatcact tacgctgagt
     1741 acttcgaaat gtccgttcgg ttggcagaag ctatgaaacg atatgggctg aatacaaatc
     1801 acagaatcgt cgtatgcagt gaaaactctc ttcaattctt tatgccggtg ttgggcgcgt
     1861 tatttatcgg agttgcagtt gcgcccgcga acgacattta taatgaacgt gaattgctca
     1921 acagtatggg catttcgcag cctaccgtgg tgttcgtttc caaaaagggg ttgcaaaaaa
     1981 ttttgaacgt gcaaaaaaag ctcccaatca tccaaaaaat tattatcatg gattctaaaa
     2041 cggattacca gggatttcag tcgatgtaca cgttcgtcac atctcatcta cctcccggtt
     2101 ttaatgaata cgattttgtg ccagagtcct tcgataggga caagacaatt gcactgatca
     2161 tgaactcctc tggatctact ggtctgccta aaggtgtcgc tctgcctcat agaactgcct
     2221 gcgtgagatt ctcgcatgcc agagatccta tttttggcaa tcaaatcatt ccggatactg
     2281 cgattttaag tgttgttcca ttccatcacg gttttggaat gtttactaca ctcggatatt
     2341 tgatatgtgg atttcgagtc gtcttaatgt atagatttga agaagagctg tttctgagga
     2401 gccttcagga ttacaagatt caaagtgcgc tgctggtgcc aaccctattc tccttcttcg
     2461 ccaaaagcac tctgattgac aaatacgatt tatctaattt acacgaaatt gcttctggtg
     2521 gcgctcccct ctctaaggaa gtcggggaag cggttgccaa gaggttccat ctgccaggta
     2581 tcaggcaagg atatgggctc actgagacta catcagctat tctgattaca cccgaggggg
     2641 atgataaacc gggcgcggtc ggtaaagttg ttccattttt tgaagcgaag gttgtggatc
     2701 tggataccgg gaaaacgctg ggcgttaatc aaagaggcga actgtgtgtg agaggtccta
     2761 tgattatgtc cggttatgta aacaatccgg aagcgaccaa cgccttgatt gacaaggatg
     2821 gatggctaca ttctggagac atagcttact gggacgaaga cgaacacttc ttcatcgttg
     2881 accgcctgaa gtctctgatt aagtacaaag gctatcaggt ggctcccgct gaattggaat
     2941 ccatcttgct ccaacacccc aacatcttcg acgcaggtgt cgcaggtctt cccgacgatg
     3001 acgccggtga acttcccgcc gccgttgttg ttttggagca cggaaagacg atgacggaaa
     3061 aagagatcgt ggattacgtc gccagtcaag taacaaccgc gaaaaagttg cgcggaggag
     3121 ttgtgtttgt ggacgaagta ccgaaaggtc ttaccggaaa actcgacgca agaaaaatca
     3181 gagagatcct cataaaggcc aagaagggcg gaaagatcgc cgtgtaactc gaggatcgtt
     3241 caaacatttg gcaataaagt ttcttaagat tgaatcctgt tgccggtctt gcgacgatta
     3301 tcatataatt tctgttgaat tacgttaagc atgtaataat taacatgtaa tgcatgacgt
     3361 tatttatgag atgggttttt atgattagag tcccgcaatt atacatttaa tacgcgatag
     3421 aaaacaaaat atagcgcgca aactaggata aattatcgcg cgcggtgtca tctatgttac
     3481 tagatcaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg
     3541 atagtactaa catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag
     3601 gctgtcccca gtgcaagtgc aggtgccaga acatttctct gagctccagc ttttgttccc
     3661 tttagtgagg gttaatttcg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     3721 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     3781 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     3841 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     3901 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     3961 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     4021 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4081 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     4141 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     4201 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     4261 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     4321 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     4381 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     4441 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     4501 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     4561 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     4621 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     4681 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4741 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     4801 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     4861 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     4921 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4981 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5041 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     5101 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     5161 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     5221 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     5281 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     5341 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     5401 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     5461 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     5521 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     5581 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     5641 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     5701 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5761 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     5821 cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt
     5881 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     5941 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     6001 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     6061 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     6121 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     6181 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     6241 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     6301 cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     6361 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     6421 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgaattgt aatacgactc
     6481 actatagggc gaattgggta caagcttgat catgccgaat tccgaacgcg ttccaacact
     6541 cgaatcccca cccgttggac caaacccggc tcattaagcg tcggttcaga tttatttcct
     6601 ttatttaaaa aaaaggaaag ggtaaaaaat agaaaattgg aaacagttaa agcccaaaat
     6661 tgtaatttac cgagaattgt aaatttacct tatagaaccg atcctcccat tggactataa
     6721 atatacctca tcaggaacat gttgggaatc ccctcgtcgc tgcgtctctc tcccgcaacc
     6781 ttcgattttc gtttattcgc atccatcgga gagagaaaac aatcaataag cgaccggatc
     6841 cgtcgacata actgcagatg acttcaaagg tgtatgatcc tgagcaggta agtttctgct
     6901 tctacctttg atatatatat aataattatc attaattagt agtaatataa tatttcaaat
     6961 atttttttca aaataaaaga atgtagtata tagcaattgc ttttctgtag tttataagtg
     7021 tgtatatttt aatttataac ttttctaata tatgaccaaa atttgttgat gtgcagagaa
     7081 aaagaatgat aactggtcca caatggtggg caagatgtaa acagatgaac gtgcttgatt
     7141 ctttcattaa ttactacgat tcagaaaagc acgctgagaa tgcagttatc tttcttcacg
     7201 gaaacgctgc atcttcatat ttgtggagac atgttgtgcc tcacattgaa ccagtggcta
     7261 ggtgtattat cccagatct
//
