LOCUS       sequence-106546-        7387 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Not gate genetic circuit containing one PhlF binding site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7387)
  TITLE     Salis Lab Plasmids
REFERENCE   2  (bases 1 to 7387)
  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"
COMMENT     Sequence Label: PhLF_N_1
FEATURES             Location/Qualifiers
     source          1..7387
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(132..149)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(266..811)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(346..365)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     883..902
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(1877..2959)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     2921..2940
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(3094..3099,3100..3115,3116..3122)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    3128..3147
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     misc_RNA        3150..3199
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     terminator      3873..3944
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3960..3987
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             4068..4079
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(4086..4088,4089..4091,4092..4805)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4131..4152)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4392..4411)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4739..4760
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4845..4862)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5015..5101
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5193..5220
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        5239..5330
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5331..5399,5400..6191)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      6233..6688
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6320..6339)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gctagcggag tgtatactgg cttactatgt tggcactgat gagggtgtca gtgaagtgct
       61 tcatgtggca ggagaaaaaa ggctgcaccg gtgcgtcagc agaatatgtg atacaggata
      121 tattccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg gcgagcggaa
      181 atggcttacg aacggggcgg agatttcctg gaagatgcca ggaagatact taacagggaa
      241 gtgagagggc cgcggcaaag ccgtttttcc ataggctccg cccccctgac aagcatcacg
      301 aaatctgacg ctcaaatcag tggtggcgaa acccgacagg actataaaga taccaggcgt
      361 ttccccctgg cggctccctc gtgcgctctc ctgttcctgc ctttcggttt accggtgtca
      421 ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga cactcagttc cgggtaggca
      481 gttcgctcca agctggactg tatgcacgaa ccccccgttc agtccgaccg ctgcgcctta
      541 tccggtaact atcgtcttga gtccaacccg gaaagacatg caaaagcacc actggcagca
      601 gccactggta attgatttag aggagttagt cttgaagtca tgcgccggtt aaggctaaac
      661 tgaaaggaca agttttggtg actgcgctcc tccaagccag ttacctcggt tcaaagagtt
      721 ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt tttcagagca
      781 agagattacg cgcagaccaa aacgatctca agaagatcat cttattaatc agataaaata
      841 tttgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata
      901 ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag
      961 gatctgctca tgtttgacag cttatcatcg atgcataatg tgcctgtcaa atggacgaag
     1021 cagggattct gcaaacccta tgctactccc tcgagccgtc aattgtctga ttcgttacca
     1081 attattaatt tttaaagtat gggcaatcaa ttgctcctgt taaaattgct ttagaaatac
     1141 tttggcagcg gtttgttgta ttgagtttca tttgcgcatt ggttaaatgg aaagtgacag
     1201 tacgctcact gcaacctaat atttttgaaa tatcccaaga gctttttcct tcgcatgccc
     1261 acgctaaaca ttctttttct cttttggtta aatcgttgtt tgatttatta tttgctatat
     1321 ttatttttcg ataattatca actagagaag gaacaattaa tggtatgttc atacacgcat
     1381 gtaaaaataa actatctata tagttgtctt tttctgaatg tgcaaaacta agcattccga
     1441 agccattgtt agccgtatga atagggaaac taaacccagt gataagacct gatgttttcg
     1501 cttctttaat tacatttgga gattttttat ttacagcatt gttttcaaat atattccaat
     1561 taattggtga atgattggag ttagaataat ctactatagg atcatatttt attaaattag
     1621 cgtcatcata atattgcctc cattttttag ggtaattatc taggattgaa atatcagatt
     1681 taaccataga atgaggataa atgatcgcga gtaaataata ttcacaatgt accattttag
     1741 tcatatcaga taagcattga ttaatatcat tattgcttct acaagcttta attttattaa
     1801 ttattctgta tgtgtcgtcg gcatttatgt ttttcatcga ggcctttcct gtgtgacccg
     1861 ctggaatagg atcttatcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     1921 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt
     1981 ttcttttcac cagtgagact ggcaacagct gattgccctt caccgcctgg ccctgagaga
     2041 gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg
     2101 ttaacggcgg gatataacat gagctatctt cggtatcgtc gtatcccact accgagatat
     2161 ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat
     2221 cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt
     2281 gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc
     2341 gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc
     2401 ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg
     2461 taccgtcctc atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa
     2521 ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg
     2581 gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac
     2641 aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg
     2701 cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg
     2761 caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt
     2821 aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg
     2881 cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt
     2941 ataacgttac tggtttcata ttcaccaccc tgaattgact ctcttccggg cgctatcatg
     3001 ccataccgcg aaaggttttg cgccattcga tggcgcgccg cttcgtcagg ccacatagct
     3061 ttcttgttct gatcggaacg atcgttggct gtgttgacaa ttaatcatcg gctcgtataa
     3121 tgtgtggaat tgtgagcgct cacaattagc tgtcaccgga tgtgctttcc ggtctgatga
     3181 gtccgtgagg acgaaacagc ctctacaaat aattttgttt aaattagtaa cataagcaag
     3241 agcctcatgg cacgtacccc gagccgtagc agcattggta gcctgcgtag tccgcatacc
     3301 cataaagcaa ttctgaccag caccattgaa atcctgaaag aatgtggtta tagcggtctg
     3361 agcattgaaa gcgttgcacg tcgtgccggt gcaagcaaac cgaccattta tcgttggtgg
     3421 accaataaag cagcactgat tgccgaagtg tatgaaaatg aaagcgaaca ggtgcgtaaa
     3481 tttccggatc tgggtagctt taaagccgat ctggattttc tgctgcgtaa tctgtggaaa
     3541 gtttggcgtg aaaccatttg tggtgaagca tttcgttgtg ttattgcaga agcacagctg
     3601 gaccctgcaa ccctgaccca gctgaaagat cagtttatgg aacgtcgtcg tgagatgccg
     3661 aaaaaactgg ttgaaaatgc cattagcaat ggtgaactgc cgaaagatac caatcgtgaa
     3721 ctgctgctgg atatgatttt tggtttttgt tggtatcgcc tgctgaccga acagctgacc
     3781 gttgaacagg atattgaaga atttaccttc ctgctgatta atggtgtttg tccgggtaca
     3841 cagcgttaac tagggcccat acccccaggc atcaaataaa acgaaaggct cagtcgaaag
     3901 actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctctactag agtcacactg
     3961 gctcaccttc gggtgggcct ttctgcgttt atacgacgta cggtggaatc tgattcgtta
     4021 ccaattgaca tgatacgaaa cgtaccgtat cgttaaggtt actagagaaa gaggagaaat
     4081 actagatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     4141 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     4201 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     4261 ccaccctcgt gaccaccttc ggctacggcc tgcaatgctt cgcccgctac cccgaccaca
     4321 tgaagctgca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     4381 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     4441 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     4501 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     4561 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     4621 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     4681 accactacct gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     4741 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     4801 agtaataagc ttggctgttt tggcggatga gagaagattt tcagcctgat acagattaaa
     4861 tcagaacgca gaagcggtct gataaaacag aatttgcctg gcggcagtag cgcggtggtc
     4921 ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta gcgccgatgg tagtgtgggg
     4981 tctccccatg cgagagtagg gaactgccag gcatcaaata aaacgaaagg ctcagtcgaa
     5041 agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga gtaggacaaa
     5101 tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc gggcaggacg
     5161 cccgccataa actgccaggc atcaaattaa gcagaaggcc atcctgacgg atggcctttt
     5221 tgcgtttcta caaactcttt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     5281 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagact atgagtattc
     5341 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     5401 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     5461 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     5521 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg
     5581 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5641 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5701 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5761 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5821 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa
     5881 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5941 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     6001 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     6061 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     6121 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     6181 agcattggta actgtcagac caagtttact catatatact ttagattgat ttacgcgccc
     6241 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     6301 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     6361 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     6421 cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc
     6481 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     6541 ttccaaactt gaacaacact caaccctatc tcgggctatt cttttgattt ataagggatt
     6601 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     6661 tttaacaaaa tattaacgtt tacaatttaa aaggatctag gtgaagatcc tttttgataa
     6721 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     6781 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     6841 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     6901 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     6961 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     7021 cctgttacca gtcaggcatt tgagaagcac acggtcacac tgcttccggt agtcaataaa
     7081 ccggtaaacc agcaatagac ataagcggct atttaacgac cctgccctga accgacgacc
     7141 gggtcgaatt tgctttcgaa tttctgccat tcatccgctt attatcactt attcaggcgt
     7201 agcaccaggc gtttaagggc accaataact gccttaaaaa aattacgccc cgccctgcca
     7261 ctcatcgcag tactgttgta attcattaag cattctgccg acatggaagc catcacagac
     7321 ggcatgatga acctgaatcg ccagcggcat cagcaccttg tcgccttgcg tataatattt
     7381 gcccatg
//
