LOCUS       sequence-355805-        4730 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ratiometric pHLuorin2 backbone (C terminal fusion).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4730)
  AUTHORS   Linders PTA, Ioannidis M, Ter Beest M, van den Bogaart G
  TITLE     Fluorescence Lifetime Imaging of pH along the Secretory Pathway.
  JOURNAL   ACS Chem Biol. 2022 Jan 21;17(1):240-251. doi: 
            10.1021/acschembio.1c00907. Epub 2022 Jan 10.
  PUBMED    35000377
REFERENCE   2  (bases 1 to 4730)
  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/acschembio.1c00907"; journalName: "ACS Chem Biol"; date: 
            "2022-01-21- 21"; volume: "17"; issue: "1"; pages: "240-251"
COMMENT     Sequence Label: pRpHLuorin2-N1
FEATURES             Location/Qualifiers
     source          1..4730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        107..410
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        411..614
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     565..585
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    637..717
                     /label=MCS
                     /note="multiple cloning site"
     primer_bind     complement(1028..1047)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    1564..1685
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1601..1620)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1655..1674
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1692..2147)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1829..1850)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2041..2060
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2174..2278
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2280..2637
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2280..2471
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2303..2323)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2488..2623
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2672..3466
                     /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             2672..3466
                     /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(2726..2745)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3336..3355
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3654..3673)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3698..3745
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4074..4662
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4563..4582
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcattagt tattaatagt
       61 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
      121 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
      181 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
      241 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
      301 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
      361 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
      421 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
      481 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
      541 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
      601 atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg actcagatct
      661 cgagctcaag cttcgaattc tgcagtcgac ggtaccgcgg gcccgggatc caccggtcgc
      721 caccatgagc aagggcgagg agctgttcac cggcgtggtg cccatcctgg tggagctgga
      781 cggcgacgtg aacggccaca agttcagcgt gagcggcgag ggcgagggcg acgccaccta
      841 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
      901 cctggtgacc accctgagct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
      961 gcagcacgac ttcttcaaga gcgccatgcc cgagggctac gtgcaggagc gcaccatctt
     1021 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     1081 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctgggcca
     1141 caagctggag tacaactaca acgagcacct ggtctacatc atggccgaca agcagaagaa
     1201 cggcaccaag gccatcttcc aggtgcacca caacatcgag gacggcggcg tgcagctggc
     1261 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     1321 ctacctgcac acccagagcg ccctgagcaa ggaccccaac gagaagcgcg accacatggt
     1381 gctgctggag ttcgtgaccg ccgccggcat cacccacggc atggacgagc tgtacaagta
     1441 aagcggccgc gactctagat cataatcagc cataccacat ttgtagaggt tttacttgct
     1501 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     1561 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     1621 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     1681 tcttaaggcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     1741 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     1801 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     1861 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     1921 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     1981 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2041 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2101 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt
     2161 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     2221 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc
     2281 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2341 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     2401 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2461 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     2521 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     2581 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat
     2641 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     2701 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     2761 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     2821 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     2881 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     2941 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     3001 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     3061 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     3121 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     3181 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     3241 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     3301 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     3361 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     3421 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     3481 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     3541 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3601 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca
     3661 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     3721 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     3781 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     3841 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     3901 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa
     3961 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4021 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4081 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4141 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     4201 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     4261 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     4321 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     4381 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     4441 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     4501 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     4561 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     4621 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     4681 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
//
