LOCUS       sequence-263434-        5174 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of an inverse-response citrate sensor variant 
            with medium binding affinity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5174)
  AUTHORS   Zhao Y, Shen Y, Wen Y, Campbell RE
  TITLE     High-Performance Intensiometric Direct- and Inverse-Response 
            Genetically Encoded Biosensors for Citrate.
  JOURNAL   ACS Cent Sci. 2020 Aug 26;6(8):1441-1450. doi: 
            10.1021/acscentsci.0c00518. Epub 2020 Jul 9.
  PUBMED    32875085
REFERENCE   2  (bases 1 to 5174)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acscentsci.0c00518"; journalName: "ACS Cent Sci"; date: 
            "2020-08-26- 26"; volume: "6"; issue: "8"; pages: "1441-1450"
COMMENT     Sequence Label: pBAD-Citroff1-R251
FEATURES             Location/Qualifiers
     source          1..5174
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             331..348
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             352..384
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(388..396,397..411)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     primer_bind     complement(472..493)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(733..752)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1470..1491
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1575..1592)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1745..1831
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1923..1950
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1969..2060
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2061..2129,2130..2921)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2279..2298)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3092..3680
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3581..3600
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3834..3851
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3866..4006
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3932..3951)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4092..4114
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4270..5148)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccat ggggggttct catcatcatc atcatcatgg tatggctagc
      361 atgactggtg gacagcaaat gggtcgggat ctgtacgacg atgacgataa ggatcctggc
      421 tcgagaatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
      481 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggtga gggcgatgcc
      541 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
      601 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
      661 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacatcca ggagcgcacc
      721 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
      781 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
      841 gggcacaagc tggagtacaa ctggcagacc gaggagcgtc tgcattatca ggtcgggcaa
      901 cgggcgctga ttcaggcgat gcagatttcg gcgatgccgg agctggtgga ggcagtgcag
      961 aaacgcgatc tcgccagaat caaagccctt atcggcccca tgcgttcgtt ctccgacgcc
     1021 acctacatca ccgtcggtga tgccagcggc cagcgcctct atcacgtcaa tcctgatgaa
     1081 atcggcaaat cgatggaagg cggcgatagc gatgaggcgt tgataaacgc taaaagctac
     1141 gtgtcagtgc gcaaaggctc gctgggatcg tcgctggccg gtaaatcgcc gattcaggac
     1201 gcaaccggca aggtgatcgg cattgtgtcg gtaggctata ccattgagca actggaaccg
     1261 aattttgtct atatcatggc cgacaagcag aagaacggca tcaaggcgaa cttccacatc
     1321 cgccacaaca tcgaggacgg cggcgtgcag ctcgcctacc actaccagca gaacaccccc
     1381 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcgtgca gtcccaactt
     1441 tcgaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     1501 gggatcactc tcggcatggt cgagctgtac aagtaaaagc ttggctgttt tggcggatga
     1561 gagaagattt tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag
     1621 aatttgcctg gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg
     1681 aaacgccgta gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag
     1741 gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt
     1801 gtcggtgaac gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa
     1861 gcaacggccc ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa
     1921 gcagaaggcc atcctgacgg atggcctttt tgcgtttcta caaactcttt tgtttatttt
     1981 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     2041 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     2101 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     2161 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     2221 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     2281 tatgtggcgc ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac
     2341 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     2401 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     2461 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     2521 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     2581 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     2641 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     2701 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     2761 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     2821 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     2881 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     2941 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     3001 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     3061 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     3121 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     3181 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     3241 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     3301 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     3361 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     3421 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     3481 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     3541 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     3601 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     3661 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     3721 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     3781 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     3841 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     3901 gtatttcaca ccgcatatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     3961 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     4021 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     4081 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     4141 gaggcagcag atcaattcgc gcgcgaaggc gaagcggcat gcataatgtg cctgtcaaat
     4201 ggacgaagca gggattctgc aaaccctatg ctactccgtc aagccgtcaa ttgtctgatt
     4261 cgttaccaat tatgacaact tgacggctac atcattcact ttttcttcac aaccggcacg
     4321 gaactcgctc gggctggccc cggtgcattt tttaaatacc cgcgagaaat agagttgatc
     4381 gtcaaaacca acattgcgac cgacggtggc gataggcatc cgggtggtgc tcaaaagcag
     4441 cttcgcctgg ctgatacgtt ggtcctcgcg ccagcttaag acgctaatcc ctaactgctg
     4501 gcggaaaaga tgtgacagac gcgacggcga caagcaaaca tgctgtgcga cgctggcgat
     4561 atcaaaattg ctgtctgcca ggtgatcgct gatgtactga caagcctcgc gtacccgatt
     4621 atccatcggt ggatggagcg actcgttaat cgcttccatg cgccgcagta acaattgctc
     4681 aagcagattt atcgccagca gctccgaata gcgcccttcc ccttgcccgg cgttaatgat
     4741 ttgcccaaac aggtcgctga aatgcggctg gtgcgcttca tccgggcgaa agaaccccgt
     4801 attggcaaat attgacggcc agttaagcca ttcatgccag taggcgcgcg gacgaaagta
     4861 aacccactgg tgataccatt cgcgagcctc cggatgacga ccgtagtgat gaatctctcc
     4921 tggcgggaac agcaaaatat cacccggtcg gcaaacaaat tctcgtccct gatttttcac
     4981 caccccctga ccgcgaatgg tgagattgag aatataacct ttcattccca gcggtcggtc
     5041 gataaaaaaa tcgagataac cgttggcctc aatcggcgtt aaacccgcca ccagatgggc
     5101 attaaacgag tatcccggca gcaggggatc attttgcgct tcagccatac ttttcatact
     5161 cccgccattc agag
//
