LOCUS       sequence-321987-        7309 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression in mammalian cells of Phosphatidic Acid preferring 
            Phospholipase A1 tagged with mEos2 to perform sptPALM.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7309)
  AUTHORS   Gormal RS, Padmanabhan P, Kasula R, Bademosi AT, Coakley S, 
            Giacomotto J, Blum A, Joensuu M, Wallis TP, Lo HP, Budnar S, Rae J, 
            Ferguson C, Bastiani M, Thomas WG, Pardon E, Steyaert J, Yap AS, 
            Goodhill GJ, Hilliard MA, Parton RG, Meunier FA
  TITLE     Modular transient nanoclustering of activated beta2-adrenergic 
            receptors revealed by single-molecule tracking of 
            conformation-specific nanobodies.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Nov 19. pii: 2007443117. doi: 
            10.1073/pnas.2007443117.
  PUBMED    33214152
REFERENCE   2  (bases 1 to 7309)
  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: "Proc Natl 
            Acad Sci U S A. 2020 Nov 19. pii: 2007443117. doi: 
            10.1073/pnas.2007443117."
COMMENT     Sequence Label: pmEos2-C1 PA-PLA1 (mEos2-PA-PLA1)
FEATURES             Location/Qualifiers
     source          1..7309
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             553..1230
                     /codon_start=1
                     /product="monomeric variant of green-to-red photoswitchable
                     fluorescent protein EosFP (McKinney et al., 2009)"
                     /label=mEos2
                     /translation="MSAIKPDMKIKLRMEGNVNGHHFVIDGDGTGKPFEGKQSMDLEVK
                     EGGPLPFAFDILTTAFHYGNRVFAKYPDNIQDYFKQSFPKGYSWERSLTFEDGGICIAR
                     NDITMEGDTFYNKVRFYGTNFPANGPVMQKKTLKWEPSTEKMYVRDGVLTGDIHMALLL
                     EGNAHYRCDFRTTYKAKEKGVKLPGYHFVDHCIEILSHDKDYNKVKLYEHAVAHSGLPD
                     NARR"
     polyA_signal    4037..4158
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4074..4093)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4128..4147
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4165..4620)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4302..4323)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4514..4533
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4647..4751
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4753..5110
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4753..4944
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4776..4796)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4961..5096
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5145..5939
                     /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             5145..5939
                     /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(5199..5218)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5809..5828
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6127..6146)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6171..6218
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6547..7135
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7036..7055
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggtcgcca ccatgagtgc gattaagcca gacatgaaga tcaaactccg tatggaaggc
      601 aacgtaaacg ggcaccactt tgtgatcgac ggagatggta caggcaagcc ttttgaggga
      661 aaacagagta tggatcttga agtcaaagag ggcggacctc tgccttttgc ctttgatatc
      721 ctgaccactg cattccatta cggcaacagg gtattcgcca aatatccaga caacatacaa
      781 gactatttta agcagtcgtt tcctaagggg tattcgtggg aacgaagctt gactttcgaa
      841 gacgggggca tttgcattgc cagaaacgac ataacaatgg aaggggacac tttctataat
      901 aaagttcgat tttatggtac caactttccc gccaatggtc cagttatgca gaagaagacg
      961 ctgaaatggg agccctccac tgagaaaatg tatgtgcgtg atggagtgct gacgggtgat
     1021 attcatatgg ctttgttgct tgaaggaaat gcccattacc gatgtgactt cagaactact
     1081 tacaaagcta aggagaaggg tgtcaagtta ccaggctacc actttgtgga ccactgcatt
     1141 gagattttaa gccatgacaa agattacaac aaggttaagc tgtatgagca tgctgttgct
     1201 cattctggat tgcctgacaa tgccagacga gagctgtaca agtccggact cagatctatg
     1261 aattacccgg gccgcgggtc cccacggagc cccgagcata acggccgagg cggcggcggc
     1321 ggcgcctggg agctgggctc agacgcgagg ccagcgttcg gcggcggcgt ctgctgcttc
     1381 gagcacctgc ccggcgggga cccggacgac ggcgacgtgc ccctggccct gctgcgcggg
     1441 gaacccgggc tgcatttggc gccgggcacc gacgaccaca accaccacct cgcgctggac
     1501 ccctgcctca gtgacgagaa ctatgacttc agctccgccg agtcgggctc ctcgctgcgc
     1561 tactacagcg agggtgagag cggcggcggc ggcagctcct tgtcgctgca cccgccgcag
     1621 cagcctccgc tggtcccgac gaactcgggg ggcggcggcg cgacaggagg gtcccccggg
     1681 gaaaggaaac gtacccggct tggcggcccg gcggcccggc accgctatga ggtagtgacg
     1741 gagctgggcc cggaggaggt acgctggttc tacaaggagg acaagaagac ctggaagccc
     1801 ttcatcggct acgactcgct ccgcatcgag ctcgccttcc ggaccctgct gcagaccacg
     1861 ggtgcccggc cccagggcgg ggaccgggac ggcgaccatg tgtgctcccc cacgggccca
     1921 gcctccagtt ccggagaaga tgacgatgag gaccgcgcct gcggcttctg ccagagtacg
     1981 acggggcacg agccggagat ggtggagctt gtgaacatcg agcctgtgtg cgtgcggggc
     2041 ggcctctacg aggtggatgt gacccaagga gagtgctacc cggtgtactg gaaccaggct
     2101 gataaaatac cagtaatgcg tggacagtgg tttattgacg gcacttggca gcctctagaa
     2161 gaggaagaaa gtaatttaat tgagcaagaa catctcaatt gttttagggg ccagcagatg
     2221 caggaaaatt tcgatattga agtgtcaaaa tccatagatg gaaaagatgc tgttcatagt
     2281 ttcaagttga gtcgaaacca tgtggactgg cacagtgtgg atgaagtata tctttatagt
     2341 gatgcaacaa catctaaaat tgcaagaaca gttacccaaa aactgggatt ttctaaagca
     2401 tcaagtagtg gtaccagact tcatagaggt tatgtagaag aagccacatt agaagacaag
     2461 ccatcacaga ctacccatat tgtatttgtt gtgcatggca ttgggcagaa aatggaccaa
     2521 ggaagaatta tcaaaaatac agctatgatg agagaagctg caagaaaaat agaagaaagg
     2581 catttttcca accatgcaac acatgttgaa tttctgcctg ttgagtggcg gtcaaaactt
     2641 actcttgatg gagacactgt tgattccatt actcctgaca aagtacgagg tttaagggat
     2701 atgctgaaca gcagtgcaat ggacataatg tattatacta gtccacttta tagagatgaa
     2761 ctagttaaag gccttcagca agagctgaat cgattgtatt cccttttctg ttctcggaat
     2821 ccagactttg aagaaaaagg gggtaaagtc tcaatagtat cacattcctt gggatgtgta
     2881 attacttatg acataatgac tggctggaat ccagttcggc tgtatgaaca gttgctgcaa
     2941 aaggaagaag agttgcctga tgaacgatgg atgagctatg aagaacgaca tcttcttgat
     3001 gaactctata taacaaaacg acggctgaag gaaatagaag aacggcttca cggattgaaa
     3061 gcatcatcta tgacacaaac acctgcctta aaatttaagg ttgagaattt cttctgtatg
     3121 ggatccccat tagcagtttt cttggcgttg cgtggcatcc gcccaggaaa tactggaagt
     3181 caagaccata ttttgcctag agagatttgt aaccggttac taaatatttt tcatcctaca
     3241 gatccagtgg cttatagatt agaaccatta atactgaaac actacagcaa catttcacct
     3301 gtccagatcc actggtacaa tacttcaaat cctttacctt atgaacatat gaagccaagc
     3361 tttctcaacc cagctaaaga acctacctca gtttcagaga atgaaggcat ttcaaccata
     3421 ccaagccctg tgacctcacc agttttgtcc cgccgacact atggagaatc tataacaaat
     3481 ataggcaaag caagcatatt aggggctgct agcattggaa agggacttgg aggaatgttg
     3541 ttctcaagat ttggacgttc atctacaaca cagtcatctg aaacatcaaa agactcaatg
     3601 gaagatgaga agaagccagt tgcctcacct tctgctacca ccgtagggac acagaccctt
     3661 ccacatagca gttctggctt cctcgattct gcattggagt tggatcacag gattgatttt
     3721 gaactcagag aaggccttgt ggagagccgc tattggtcag ctgtcacgtc gcatactgcc
     3781 tattggtcat ccttggatgt tgcccttttt cttttaacct tcatgtataa acatgagcac
     3841 gatgatgatg caaaacccaa tttagatcca atctgagaat tctgcagtcg acggtaccgc
     3901 gggcccggga tccaccggat ctagataact gatcataatc agccatacca catttgtaga
     3961 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     4021 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4081 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4141 actcatcaat gtatcttaac gcgtaaattg taagcgttaa tattttgtta aaattcgcgt
     4201 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     4261 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     4321 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     4381 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     4441 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     4501 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     4561 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     4621 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     4681 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     4741 aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg
     4801 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     4861 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4921 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     4981 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     5041 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     5101 cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     5161 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     5221 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     5281 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc
     5341 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     5401 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     5461 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     5521 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     5581 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     5641 cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg
     5701 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     5761 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     5821 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     5881 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     5941 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     6001 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     6061 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag
     6121 gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa
     6181 aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     6241 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     6301 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     6361 cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa
     6421 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     6481 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     6541 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     6601 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     6661 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     6721 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     6781 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     6841 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     6901 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     6961 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     7021 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     7081 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     7141 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     7201 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat
     7261 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccg
//
