LOCUS       sequence-484658-        4534 bp    DNA     circular SYN 26-AUG-2025
DEFINITION  Expression Plasmid containing a LacI repressed sd2 AraC R5 PANCE 
            variant cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4534)
  TITLE     Phage-Assisted Evolution of Allosteric Protein Switches
REFERENCE   2  (bases 1 to 4534)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 26, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pDest_PANCEd_AraC_sd2_R5_C1
FEATURES             Location/Qualifiers
     source          1..4534
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     154..171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(379..1461)
                     /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     1423..1442
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1462..1539)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(1584..1589,1590..1606,1607..1613)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1621..1637
                     /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)."
     primer_bind     complement(2584..2601)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2754..2840
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2932..2959
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(2979..3073)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3094..3753)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LLSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3668..3687
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(3754..3856)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      3946..4534
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4435..4454
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      121 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      181 gagcgccacg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc
      241 ctcaacgaca ggagcacgat catgctagtc atgccccgcg cccaccggaa ggagctgact
      301 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
      361 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
      421 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
      481 ttttcttttc accagtgaca cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
      541 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
      601 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
      661 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
      721 atcgttggca accagcatcg cagtgggaac gataccctca ttcagcattt gcatggtttg
      781 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
      841 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
      901 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
      961 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     1021 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     1081 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     1141 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     1201 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     1261 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     1321 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     1381 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     1441 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     1501 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     1561 tatgcgactc cgatcccgca aatttgacaa ttaatcatcc ggctcgtata atgtgtggaa
     1621 ttgtgagcgg ataacaattg gagtaaaaaa aaaggaaaat gtccgaggaa gcgcagaacg
     1681 atccgctgct gccgggctat agctttaacg cgcatctggt ggcgggcctg accccgattg
     1741 aagcgaacgg ctatctggat ttttttattg atcgcccgct gggcatgaaa ggctatattc
     1801 tgaacctgac cattcgcggc cagggcgtgg tgaaaaacca gggccgcgaa tttgtgtgcc
     1861 gcccgggcga tattctgctg tttccgccgg gcgaaattca tcattatggc cgccatccgg
     1921 aagcgcgcga atggtatcat cagtgggtgt attttcgccc gcgcgcgtat tggcatgaat
     1981 ggctgaactg gccgagcatt tttgcgaaca ccggcttttt tcgcccggat gaagcgcatc
     2041 agccgcattt tagcgatctg tttggccaga ttattaacgc gggccagggc gaaggccgct
     2101 atagcgaact gctggcgatt aacctgatag aacagctgct gctgcgccgc atggaagcga
     2161 ttaacgaaag cctgcatccg ccgatggata accgcgtgcg cgaagcgtgc cagtatatta
     2221 gcgatcatct ggcggatagc aactttgata ttgcgagcgt ggcgcagcat gtgtgcctga
     2281 gcccgagccg cctgagccat ctgtttcgcc agcagctggg cattagcgtg ctgagctggc
     2341 gcgaagatca gcgcattagc caggcgaaac tgctgctgag caccacccgc atgccgattg
     2401 cgaccgtggg ccacaacgtg ggctttgatg atcagctgta ttttagccgc gtgtttaaaa
     2461 aatgcaccgg cgcgagcccg agcgaatttc gcgcgggctg cgaagaaaaa gtgaacgatg
     2521 tggcggtgaa actgagcggg taaggctaat ggagattttc aacatgggcg agaagatttt
     2581 cagcctgata cagattaaat cagaacgcag aagcggtctg ataaaacaga atttgcctgg
     2641 cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag
     2701 cgccgatggt agtgtggggt atccccatgc gagagtaggg aactgccagg catcaaataa
     2761 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     2821 ctctcctgag taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg
     2881 gagggtggcg ggcaggacgc ccgccataaa ctgccaggca tcaaattaag cagaaggcca
     2941 tcctgacgga tggccttttc tgaaaggagg aactatatat tctcaccaat aaaaaacgcc
     3001 cggcggcaac cgagcgttct gaacaaatcc agatggagtt ctgaggtcat tactggatct
     3061 atcaacagga gtccaagcga gctcgatatc aaattacgcc ccgccctgcc actcatcgca
     3121 gtactgttgt aattcattaa gcattctgcc gacatggaag ccatcacaaa cggcatgatg
     3181 aacctgaatc gccagcggca tcagcacctt gtcgccttgc gtataatatt tgcccatggt
     3241 gaaaacgggg gcgaagaagt tgtccatatt ggccacgttt aaatcaaaac tggtgaaact
     3301 cacccaggga ttggctgaaa cgaaaaacat attctcaata aaccctttag ggaaataggc
     3361 caggttttca ccgtaacacg ccacatcttg cgaatatatg tgtagaaact gccggaaatc
     3421 gtcgtggtat tcactcaaga gcgatgaaaa cgtttcagtt tgctcatgga aaacggtgta
     3481 acaagggtga acactatccc atatcaccag ctcaccgtct ttcattgcca tacgaaattc
     3541 cggatgagca ttcatcaggc gggcaagaat gtgaataaag gccggataaa acttgtgctt
     3601 atttttcttt acggtcttta aaaaggccgt aatatccagc tgaacggtct ggttataggt
     3661 acattgagca actgactgaa atgcctcaaa atgttcttta cgatgccatt gggatatatc
     3721 aacggtggta tatccagtga tttttttctc cattttagct tccttagctc ctgaaaatct
     3781 cgataactaa aaaaatacgc ccggtagtga tcttatttca ttatggtgaa agttggaacc
     3841 tcttacgtgc cgatcaacag ttttattgtt catgaccaaa atcccttaac gtgagttttc
     3901 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     3961 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4021 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     4081 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     4141 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4201 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4261 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4321 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4381 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4441 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4501 gtgatgctcg tcaggggggc ggagcctatg gaaa
//
