LOCUS       sequence-209471-        5784 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for the expression of the inactive mutant of RINS1 sensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5784)
  AUTHORS   Schifferer M, Yushchenko DA, Stein F, Bolbat A, Schultz C
  TITLE     A Ratiometric Sensor for Imaging Insulin Secretion in Single beta 
            Cells.
  JOURNAL   Cell Chem Biol. 2017 Apr 20;24(4):525-531.e4. doi: 
            10.1016/j.chembiol.2017.03.001. Epub 2017 Mar 30.
  PUBMED    28366620
REFERENCE   2  (bases 1 to 5784)
  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: 
            "10.1016/j.chembiol.2017.03.001"; journalName: "Cell Chem Biol"; 
            date: "2017-04-20- 20"; volume: "24"; issue: "4"; pages: 
            "525-531.e4"
COMMENT     Sequence Label: RINS1(mut)
FEATURES             Location/Qualifiers
     source          1..5784
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     sig_peptide     633..704
                     /codon_start=1
                     /product="signal sequence of human preproinsulin"
                     /label=insulin signal sequence
                     /translation="MALWMRLLPLLALLALWGPDPAAA"
     CDS             849..1556
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(994..1012)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1262..1281
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2572..2693
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2609..2628)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2663..2682
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2700..3155)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2837..2858)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3049..3068
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3182..3286
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3288..3645
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3288..3479
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3311..3331)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3496..3631
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3680..4474
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3680..4474
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3734..3753)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4344..4363
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4662..4681)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4706..4753
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5082..5670
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5571..5590
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttccg ccatggccct gtggatgcgc ctcctgcccc
      661 tgctggcgct gctggccctc tggggacctg acccagccgc agcctttgtg aaccaacacc
      721 tgtgcggctc acacctggtg gaagctctct acctagtgtg cggggaacga ggcttcttct
      781 acacacccaa gaccagccgg gaggcagagg acctgcaggt ggggcaggtg gagctgggcg
      841 ggggccctat ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc
      901 gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg
      961 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc
     1021 ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg
     1081 tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt
     1141 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     1201 tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg
     1261 gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg
     1321 aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact
     1381 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct
     1441 acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac
     1501 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaaggctg
     1561 ctgctgcggg ccctggtgca ggcagcctgc agcccttggc cctggagggg tccctgcaga
     1621 agagtggcat tgtggaacaa tgctgtacca gcatctgctc cctctaccag ctggagaact
     1681 actgcaactt ggatccaccg gtcgccaccg gtgcaggagc tatgagcaag ggcgaggagc
     1741 tgttcaccgg cgtggtgccc atcctggtgg agctggacgg cgacgtgaac ggccacaagt
     1801 tcagcgtgcg cggcgagggc gagggcgacg ccaccaacgg caagctgacc ctgaagttca
     1861 tctgcaccac cggcaagctg cccgtgccct ggcccaccct ggtgaccacc ctgacctacg
     1921 gcgtgcagtg cttcagccgc taccccgacc acatgaagcg ccacgacttc ttcaagagcg
     1981 ccatgcccga gggctacgtg caggagcgca ccatcagctt caaggacgac ggcacctaca
     2041 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     2101 gcatcgactt caaggaggac ggcaacatcc tgggccacaa gctggagtac aacttcaaca
     2161 gccacaacgt gtacatcacc gccgacaagc agaagaacgg catcaaggcc aacttcaaga
     2221 tccgccacaa cgtggaggac ggcagcgtgc agctggccga ccactaccag cagaacaccc
     2281 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagagccgcc
     2341 tgagcaagga ccccaacgag aagcgcgacc acatggtgct gctggagttc gtgaccgccg
     2401 ccggcatcac ccacggcatg gacgagctgt acaagggcag ctaagtcgac gcggccgcga
     2461 ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc
     2521 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
     2581 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     2641 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta
     2701 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     2761 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     2821 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     2881 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     2941 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     3001 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     3061 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     3121 acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt
     3181 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     3241 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag
     3301 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3361 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3421 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3481 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3541 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     3601 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac
     3661 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     3721 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     3781 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     3841 cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg
     3901 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     3961 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     4021 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     4081 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     4141 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     4201 caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     4261 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     4321 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     4381 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     4441 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     4501 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     4561 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     4621 ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca
     4681 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt
     4741 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca
     4801 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca
     4861 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc
     4921 tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     4981 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     5041 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     5101 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     5161 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     5221 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     5281 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     5341 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     5401 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     5461 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     5521 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     5581 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     5641 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     5701 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     5761 gataaccgta ttaccgccat gcat
//
