LOCUS       sequence-464951-        4429 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of glutamate sensor with improved sensitivity 
            and slow deactivation kinetics (4s = slow).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4429)
  AUTHORS   Aggarwal A, Negrean A, Chen Y, Iyer R, Reep D, Liu A, Palutla A, Xie
            ME, MacLennan BJ, Hagihara KM, Kinsey LW, Sun JL, Yao P, Zheng J, 
            Tsang A, Tsegaye G, Zhang Y, Patel RH, Arthur BJ, Hiblot J, Leippe 
            P, Tarnawski M, Marvin JS, Vevea JD, Turaga SC, Tebo AG, Carandini 
            M, Rossi LF, Kleinfeld D, Konnerth A, Svoboda K, Turner GC, Hasseman
            J, Podgorski K
  TITLE     Glutamate indicators with increased sensitivity and tailored 
            deactivation rates
  JOURNAL   bioRxiv 2025.03.20.643984
REFERENCE   2  (bases 1 to 4429)
  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"; journalName: "bioRxiv 
            2025.03.20.643984"
COMMENT     Sequence Label: pRSET-iGluSnFR4s
FEATURES             Location/Qualifiers
     source          1..4429
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             112..129
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             133..165
                     /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(169..177,178..192)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     primer_bind     1167..1188
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1291..1312)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1830..1848)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1844..1891
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1988..2443
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2075..2094)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2285..2306
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2469..2573
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2574..2642,2643..3434)
                     /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     complement(2792..2811)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3605..4193
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4094..4113
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4347..4364
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcggggttc tcatcatcat
      121 catcatcatg gtatggctag catgactggt ggacagcaaa tgggtcggga tctgtacgac
      181 gatgacgata aggatcgatg gggatccgcc gcaggcagca cgctggacaa aatcgccaaa
      241 aacggtgtga ttgtcgtcgg tcaccgtgac tcttcagtgc ctttctctta tcaagacagt
      301 cagcacaaag tggtgggtta ctcgcaggaa tactccaacg ccattgttga agcagtgaaa
      361 aagaaactca acaaaccgga cttgcaggta aaactgattc cgattacttc acaaaaccgt
      421 attccactac tgcaaaacgg cacttacgat tttgaatgtg gttctaccac caacaacgtc
      481 gaacgccaaa aattcgcggc tttctctgac actattttcg tggtcggtac gcgcctgttg
      541 accaaaaagg gtggcgatat caaagatttt gccaacctga aagacaaagc cgtagtcgtc
      601 acttccggca ctacctctga agttttgctc aacaaactga atgaagagca aaaaatgaat
      661 atgcgcatca tcagcgccaa agatcacggt gactctttcc gcaccctgga aagcggtcgt
      721 gccgttgcct ttatgatgga tgactcgctg ctggccggtg aacgtgcgaa agcgaagaaa
      781 ccagacgact gggaaatcgt cggcaagccg cagtctcagg aggcctacgg ttgtatgttg
      841 cgtaaagatg atccgcagtt caaaaagctg atggatgaca ccatcgctca ggtgcagacc
      901 tccggtgaag cggaaaaatg gtttgataag tggttcaaaa atccaattct ggtgaaccac
      961 acggtctata tcaccgccga caagcagaag aacggcatca aggcgaactt cttcatccac
     1021 cacaacgtgg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     1081 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gctatcagtc cgtgctgagc
     1141 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccaccggg
     1201 aactctctcg gcatggacga gctgtacaag ggcggtaccg gagggagcat gagcaagggc
     1261 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     1321 cacaagttca gcgtgcgcgg cgagggcgag ggcgatgcca ccaacggcaa gctaaccctg
     1381 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     1441 agctacggcg tgcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc
     1501 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcagcttcaa ggacgacgga
     1561 acctacaaga cccgcgccga ggtgaagtta gagggcgaca ccctggtgaa ccgcatcgag
     1621 ctaaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     1681 tttaacaacc ccctggtcat gaatttcgaa ctgtcagacg aaatgaaagc actgttcaaa
     1741 gaaccgaatg acaaggcact gaaataagaa ttcgaagctt gatccggctg ctaacaaagc
     1801 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1861 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggatctgg
     1921 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     1981 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2041 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2101 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2161 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2221 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2281 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2341 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2401 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     2461 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2521 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2581 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2641 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2701 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     2761 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     2821 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     2881 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     2941 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     3001 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     3061 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     3121 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3181 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3241 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3301 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3361 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3421 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3481 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3541 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3601 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3661 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3721 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     3781 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3841 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3901 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3961 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4021 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4081 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4141 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4201 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4261 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4321 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4381 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcag
//
