LOCUS       sequence-355810-        4959 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Trans-Golgi expression of ratiometric pHLuorin2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4959)
  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 4959)
  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: GalT-RpHLuorin2
FEATURES             Location/Qualifiers
     source          1..4959
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        108..411
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        412..615
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     566..586
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     complement(1258..1277)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    1794..1915
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1831..1850)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1885..1904
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1922..2377)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2059..2080)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2271..2290
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2404..2508
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2510..2867
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2510..2701
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2533..2553)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2718..2853
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2902..3696
                     /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             2902..3696
                     /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(2956..2975)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3566..3585
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3884..3903)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3928..3975
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4304..4892
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4793..4812
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag ttattaatag
       61 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
      121 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      181 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      241 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
      301 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
      361 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      421 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      481 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      541 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      601 tatataagca gagctggttt agtgaaccgt cagatccgct agcgctaccg gactcagatc
      661 tcgagctcaa gcttcgaatt catgaggctt cgggagccgc tcctgagcgg cagcgccgcg
      721 atgccaggcg cgtccctaca gcgggcctgc cgcctgctcg tggccgtctg cgctctgcac
      781 cttggcgtca ccctcgttta ctacctggct ggccgcgacc tgagccgcct gccccaactg
      841 gtcggagtct ccacaccgct gcagggcggc tcgaacagtg ccgccgccat cgggcagtcc
      901 tccggggagc tccggaccgg aggggccgtg gatccaggcg gcagcggcgg cagcgctagc
      961 aagggcgagg agctgttcac cggcgtggtg cccatcctgg tggagctgga cggcgacgtg
     1021 aacggccaca agttcagcgt gagcggcgag ggcgagggcg acgccaccta cggcaagctg
     1081 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctggtgacc
     1141 accctgagct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
     1201 ttcttcaaga gcgccatgcc cgagggctac gtgcaggagc gcaccatctt cttcaaggac
     1261 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     1321 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctgggcca caagctggag
     1381 tacaactaca acgagcacct ggtctacatc atggccgaca agcagaagaa cggcaccaag
     1441 gccatcttcc aggtgcacca caacatcgag gacggcggcg tgcagctggc cgaccactac
     1501 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgcac
     1561 acccagagcg ccctgagcaa ggaccccaac gagaagcgcg accacatggt gctgctggag
     1621 ttcgtgaccg ccgccggcat cacccacggc atggacgagc tgtacaagta aagcggccgc
     1681 gactctagat cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc
     1741 tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt
     1801 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     1861 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg
     1921 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     1981 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
     2041 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
     2101 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     2161 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     2221 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     2281 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     2341 acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat
     2401 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     2461 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa
     2521 agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     2581 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     2641 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     2701 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     2761 atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat
     2821 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag
     2881 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     2941 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     3001 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     3061 tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg
     3121 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     3181 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     3241 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     3301 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     3361 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     3421 ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     3481 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     3541 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     3601 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     3661 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     3721 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     3781 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     3841 atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga
     3901 caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg
     3961 ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc
     4021 caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc
     4081 caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag
     4141 cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     4201 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     4261 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     4321 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     4381 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     4441 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     4501 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     4561 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     4621 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     4681 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     4741 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     4801 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     4861 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     4921 tcctggcctt ttgctggcct tttgctcaca tgttctttc
//
