LOCUS       sequence-374871-        4784 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmids for pH sensitive fluorescent protein 
            Lime.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4784)
  AUTHORS   Shen Y, Wen Y, Sposini S, Vishwanath AA, Abdelfattah AS, Schreiter 
            ER, Lemieux MJ, de Juan-Sanz J, Perrais D, Campbell RE
  TITLE     Rational Engineering of an Improved Genetically Encoded pH Sensor 
            Based on Superecliptic pHluorin.
  JOURNAL   ACS Sens. 2023 Aug 25;8(8):3014-3022. doi: 
            10.1021/acssensors.3c00484. Epub 2023 Jul 23.
  PUBMED    37481776
REFERENCE   2  (bases 1 to 4784)
  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: 
            "10.1021/acssensors.3c00484"; journalName: "ACS Sens"; date: 
            "2023-08-25- 25"; volume: "8"; issue: "8"; pages: "3014-3022"
COMMENT     Sequence Label: pBAD-Lime
FEATURES             Location/Qualifiers
     source          1..4784
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      175..763
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     664..683
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     917..934
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    949..1089
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1015..1034)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1175..1197
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1353..2231)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        2258..2542
                     /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     2465..2484
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             2588..2605
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2609..2641
                     /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(2645..2653,2654..2668)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     primer_bind     complement(3442..3459)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3612..3698
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3790..3817
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        3836..3927
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3928..3996,3997..4784,1..4)
                     /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(4146..4165)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
       61 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
      121 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
      181 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa caaccaccgc taccagcggt
      241 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
      301 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
      361 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
      421 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
      481 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
      541 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
      601 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
      661 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
      721 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
      781 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
      841 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
      901 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
      961 ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc tcagtacaat
     1021 ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc
     1081 atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     1141 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     1201 tcaccgtcat caccgaaacg cgcgaggcag cagatcaatt cgcgcgcgaa ggcgaagcgg
     1261 catgcataat gtgcctgtca aatggacgaa gcagggattc tgcaaaccct atgctactcc
     1321 gtcaagccgt caattgtctg attcgttacc aattatgaca acttgacggc tacatcattc
     1381 actttttctt cacaaccggc acggaactcg ctcgggctgg ccccggtgca ttttttaaat
     1441 acccgcgaga aatagagttg atcgtcaaaa ccaacattgc gaccgacggt ggcgataggc
     1501 atccgggtgg tgctcaaaag cagcttcgcc tggctgatac gttggtcctc gcgccagctt
     1561 aagacgctaa tccctaactg ctggcggaaa agatgtgaca gacgcgacgg cgacaagcaa
     1621 acatgctgtg cgacgctggc gatatcaaaa ttgctgtctg ccaggtgatc gctgatgtac
     1681 tgacaagcct cgcgtacccg attatccatc ggtggatgga gcgactcgtt aatcgcttcc
     1741 atgcgccgca gtaacaattg ctcaagcaga tttatcgcca gcagctccga atagcgccct
     1801 tccccttgcc cggcgttaat gatttgccca aacaggtcgc tgaaatgcgg ctggtgcgct
     1861 tcatccgggc gaaagaaccc cgtattggca aatattgacg gccagttaag ccattcatgc
     1921 cagtaggcgc gcggacgaaa gtaaacccac tggtgatacc attcgcgagc ctccggatga
     1981 cgaccgtagt gatgaatctc tcctggcggg aacagcaaaa tatcacccgg tcggcaaaca
     2041 aattctcgtc cctgattttt caccaccccc tgaccgcgaa tggtgagatt gagaatataa
     2101 cctttcattc ccagcggtcg gtcgataaaa aaatcgagat aaccgttggc ctcaatcggc
     2161 gttaaacccg ccaccagatg ggcattaaac gagtatcccg gcagcagggg atcattttgc
     2221 gcttcagcca tacttttcat actcccgcca ttcagagaag aaaccaattg tccatattgc
     2281 atcagacatt gccgtcactg cgtcttttac tggctcttct cgctaaccaa accggtaacc
     2341 ccgcttatta aaagcattct gtaacaaagc gggaccaaag ccatgacaaa aacgcgtaac
     2401 aaaagtgtct ataatcacgg cagaaaagtc cacattgatt atttgcacgg cgtcacactt
     2461 tgctatgcca tagcattttt atccataaga ttagcggatc ctacctgacg ctttttatcg
     2521 caactctcta ctgtttctcc atacccgttt tttgggctaa caggaggaat taaccatggg
     2581 gggttctcat catcatcatc atcatggtat ggctagcatg actggtggac agcaaatggg
     2641 tcgggatctg tacgacgatg acgataagga tccgagctcg agaatggtga gtaaaggaga
     2701 agaacttttc actggagtgg tgcctatcct ggtggaactg gatggcgacg tgaacggcca
     2761 caagttcagc gttagaggcg aaggcgaagg ggatgccaca aacggcaagc tgaccctgaa
     2821 gttcatcagc accacaggca agctgcccgt gccttggcct acactggtca caacactgac
     2881 ctacggcgtg cagagcttca gcagataccc cgaccacatg aagcggcacg atttcttcaa
     2941 gagcgccatg ccagagggct acgtgcaaga gagaaccatc agcttcaagg acgacggcac
     3001 ctacaagacc agagccgaag tgaagttcga gggcgacacc ctggtcaacc ggatcgagct
     3061 gaagggcatc gacttcaaag aggacggcaa catcctgggc cacaaacttg agtacaactt
     3121 caacgaccac caggtctaca tcaccgccga caagcagaag aacggcatca aggccaactt
     3181 caagatccgg cacaacgtgg aagatggcag cgtgcagctg gccgatcact accagcagaa
     3241 cacacccatc ggagatggcc ctgtgctgct gcccgataac cactacctgt tcaccaccag
     3301 cacactgagc aaggacccca acgagaagcg ggaccacatg gtgctgctgg aatttgtgac
     3361 cgccgccggc attacacatg gcatggatga actatacaaa tgaaagcttg gctgttttgg
     3421 cggatgagag aagattttca gcctgataca gattaaatca gaacgcagaa gcggtctgat
     3481 aaaacagaat ttgcctggcg gcagtagcgc ggtggtccca cctgacccca tgccgaactc
     3541 agaagtgaaa cgccgtagcg ccgatggtag tgtggggtct ccccatgcga gagtagggaa
     3601 ctgccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct
     3661 gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgggagcg gatttgaacg
     3721 ttgcgaagca acggcccgga gggtggcggg caggacgccc gccataaact gccaggcatc
     3781 aaattaagca gaaggccatc ctgacggatg gcctttttgc gtttctacaa actcttttgt
     3841 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     3901 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     3961 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     4021 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     4081 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     4141 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
     4201 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     4261 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     4321 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     4381 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     4441 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     4501 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     4561 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     4621 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     4681 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     4741 aatagacaga tcgctgagat aggtgcctca ctgattaagc attg
//
