LOCUS       sequence-224816-        7741 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Reporter plasmid for measuring protease activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7741)
  AUTHORS   Gao XJ, Chong LS, Kim MS, Elowitz MB
  TITLE     Programmable protein circuits in living cells.
  JOURNAL   Science. 2018 Sep 21;361(6408):1252-1258. doi: 
            10.1126/science.aat5062.
  PUBMED    30237357
REFERENCE   2  (bases 1 to 7741)
  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"; doi: "10.1126/science"; 
            journalName: "Science"; date: "2018-09-21- 21"; volume: "361"; 
            issue: "6408"; pages: "1252-1258"
COMMENT     Sequence Label: PB-CMV-TO-Casp3te-cx
FEATURES             Location/Qualifiers
     source          1..7741
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(268..289)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     480..499
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     713..735
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     727..744
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   874..936
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        1409..1788
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1789..1992
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1943..1963
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    1994..2012
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2015..2033
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2043..2067
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2159..2168
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2693..2713
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(3121..3138)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3127..3334
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3367..3387)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3372..3701
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3552..3687
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3614..3633
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3749..3796
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3815..4189
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     primer_bind     complement(4249..4269)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    4311..4432
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4348..4367)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4402..4421
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   5195..5229
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(5720..5736)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5720..5736)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5733..5755)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5744..5760
                     /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(5768..5774,5775..5792,5793..5798))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5813..5834
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5951..5968)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6122..6710)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6202..6221)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6881..7672,7673..7741))
                     /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     7504..7523
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tgagggacag ccccccccca aagcccccag
     1141 ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc
     1201 cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg
     1261 ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga
     1321 cacctggggg gatacgggga aaaggcctcc acggccacta gtaggcgttt tgcgctgctt
     1381 cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa
     1441 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
     1501 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1561 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1621 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     1681 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     1741 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
     1801 tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
     1861 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     1921 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     1981 ataagcagag ctctccctat cagtgataga gatctcccta tcagtgatag agatcgtcga
     2041 cgagctcgtt tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc
     2101 catagaagac accgggaccg atccagcctc cggactctag cgtttaaact taagcttcgc
     2161 caccatggga tccgaaaaca ctgagaacag cgtcgatagc aagtccatta agaacctcga
     2221 acctaaaatc atacatggca gtgagagcat ggatagcggc ataagccttg acaactccta
     2281 taaaatggac tacccggaga tgggtctgtg cataataatc aacaacaaga acttccacaa
     2341 atccacgggg atgacaagta ggtctggaac cgatgtcgat gcggcaaact tgcgcgagac
     2401 ctttagaaac ctcaagtacg aggtacgcaa taaaaacgat ctcactcgcg aggaaatagt
     2461 agagctgatg cgagatgtca gtaaggaaga tcattccaag agatcctcct tcgtctgcgt
     2521 cttgttgagt cacggagagg aagggatcat attcggtacg aacggcccag tggatttgaa
     2581 aaagataacc aatttcttta gaggtgaccg gtgccgctct ctcacaggaa agccaaaact
     2641 ctttattatc caagcatgta ggggtacgga actggattgt ggaatcgaaa ctgagaactt
     2701 gtacttccag tccggggtag atgacgacat ggcctgccat aaaatccctg tcgaggccga
     2761 ttttctctac gcctactcta ctgcgcctgg gtattacagc tggagaaaca gcaaggatgg
     2821 ttcctggttt atacaaagcc tgtgtgctat gctcaaacag tatgcggata agctggagtt
     2881 catgcatatc cttaccagag taaatcgcaa agtggccaca gagtttgagt cattttcttt
     2941 tgacgcaaca tttcacgcca agaagcagat tccttgtata gtatctatgc tgacgaaaga
     3001 gctgtacttc tatcacagcg gcggtggagg acggagcaaa ctgaatccac ccgatgagtc
     3061 tggacccggt tgtatgtcat gtaaatgtgt gttgagctag cgaattcgaa tttaaatcgg
     3121 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     3181 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     3241 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     3301 aggattggga agacaatagc aggcatgctg gggacgcggc cgcaatttaa cgcgaattaa
     3361 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3421 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     3481 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3541 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3601 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3661 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3721 tatccatttt cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg
     3781 catagtataa tacgacaagg tgaggaacta aaccatggcc aagttgacca gtgccgttcc
     3841 ggtgctcacc gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc ggctcgggtt
     3901 ctcccgggac ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg tgaccctgtt
     3961 catcagcgcg gtccaggacc aggtggtgcc ggacaacacc ctggcctggg tgtgggtgcg
     4021 cggcctggac gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact tccgggacgc
     4081 ctccgggccg gccatgaccg agatcggcga gcagccgtgg gggcgggagt tcgccctgcg
     4141 cgacccggcc ggcaactgcg tgcacttcgt ggccgaggag caggactgag tcgacaatca
     4201 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     4261 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gcgttaacta aacttgttta
     4321 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4381 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     4441 ggaattgact caaatgatgt caattagtct atcagaagct catctggtct cccttccggg
     4501 ggacaagaca tccctgttta atatttaaac agcagtgttc ccaaactggg ttcttatatc
     4561 ccttgctctg gtcaaccagg ttgcagggtt tcctgtcctc acaggaacga agtccctaaa
     4621 gaaacagtgg cagccaggtt tagccccgga attgactgga ttcctttttt agggcccatt
     4681 ggtatggctt tttccccgta tccccccagg tgtctgcagg ctcaaagagc agcgagaagc
     4741 gttcagagga aagcgatccc gtgccacctt ccccgtgccc gggctgtccc cgcacgctgc
     4801 cggctcgggg atgcgggggg agcgccggac cggagcggag ccccgggcgg ctcgctgctg
     4861 ccccctagcg ggggagggac gtaattacat ccctgggggc tttggggggg ggctgtccct
     4921 gatatctata acaagaaaat atatatataa taagttatca cgtaagtaga acatgaaata
     4981 acaatataat tatcgtatga gttaaatctt aaaagtcacg taaaagataa tcatgcgtca
     5041 ttttgactca cgcggtcgtt atagttcaaa atcagtgaca cttaccgcat tgacaagcac
     5101 gcctcacggg agctccaagc ggcgactgag atgtcctaaa tgcacagcga cggattcgcg
     5161 ctatttagaa agagagagca atatttcaag aatgcatgcg tcaattttac gcagactatc
     5221 tttctagggt taatctagct gcatcaggat catatcgtcg ggtctttttt ccggctcagt
     5281 catcgcccaa gctggcgcta tctgggcatc ggggaggaag aagcccgtgc cttttcccgc
     5341 gaggttgaag cggcatggaa agagtttgcc gaggatgact gctgctgcat tgacgttgag
     5401 cgaaaacgca cgtttaccat gatgattcgg gaaggtgtgg ccatgcacgc ctttaacggt
     5461 gaactgttcg ttcaggccac ctgggatacc agttcgtcgc ggcttttccg gacacagttc
     5521 cggatggtca gcccgaagcg catcagcaac ccgaacaata ccggcgacag ccggaactgc
     5581 cgtgccggtg tgcagattaa tgacagcggt gcggcgctgg gatattacgt cagcgaggac
     5641 gggtatcctg gctggatgcc gcagaaatgg acatggatac cccgtgagtt acccggcggg
     5701 cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     5761 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     5821 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     5881 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     5941 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     6001 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     6061 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     6121 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     6181 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     6241 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     6301 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     6361 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     6421 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     6481 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     6541 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     6601 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     6661 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     6721 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     6781 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     6841 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6901 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6961 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     7021 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     7081 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     7141 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     7201 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     7261 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     7321 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     7381 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     7441 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     7501 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     7561 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     7621 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     7681 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     7741 t
//
