LOCUS       sequence-399477-        5861 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses fluorescent protein-tagged enzyme.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5861)
  AUTHORS   Wills RC, Doyle CP, Zewe JP, Pacheco J, Hansen SD, Hammond GRV
  TITLE     A novel homeostatic mechanism tunes PI(4,5)P2-dependent signaling at
            the plasma membrane.
  JOURNAL   J Cell Sci. 2023 Aug 3:jcs.261494. doi: 10.1242/jcs.261494.
  PUBMED    37534432
REFERENCE   2  (bases 1 to 5861)
  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: "J Cell Sci.
            2023 Aug 3:jcs.261494. doi: 10.1242/jcs.261494."
COMMENT     Sequence Label: mCherry-Mss4-Kina
FEATURES             Location/Qualifiers
     source          1..5861
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             127..921
                     /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             127..921
                     /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(181..200)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     791..810
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1109..1128)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1153..1200
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1529..2117
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2018..2037
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2292..2595
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2596..2799
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2750..2770
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2838..2847
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2844..3551
                     /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(2989..3007)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3257..3276
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    4880..5001
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4917..4936)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4971..4990
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5008..5463)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5145..5166)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5357..5376
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5490..5594
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(5596..5861,1..92)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5596..5787
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5619..5639)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(5804..5861,1..78)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
       61 gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg
      121 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
      181 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
      241 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
      301 gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
      361 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
      421 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
      481 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
      541 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
      601 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg
      661 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
      721 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
      781 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
      841 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
      901 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
      961 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     1021 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     1081 tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa
     1141 cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc
     1201 ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt
     1261 ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag
     1321 gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat
     1381 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     1441 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     1501 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     1561 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     1621 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     1681 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     1741 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     1801 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     1861 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     1921 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     1981 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     2041 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     2101 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     2161 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     2221 ccgccatgca ttagttatta atagtaatca attacggggt cattagttca tagcccatat
     2281 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     2341 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     2401 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     2461 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     2521 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     2581 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     2641 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     2701 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     2761 ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc
     2821 cgctagcgct accggtcgcc accatggtga gcaagggcga ggaggataac atggccatca
     2881 tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg
     2941 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
     3001 tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg
     3061 gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc
     3121 ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga
     3181 cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca
     3241 acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg
     3301 agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga
     3361 aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc
     3421 agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact
     3481 acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg
     3541 agctgtacaa gtccggactc agatctcgag ctcaagcttc gaattctgca gtcgacggta
     3601 ccgcgaacaa ggtctcagag ggtcacgtga atttcatcat tgcctacaac atgctcactg
     3661 gcataagagt tgccgtcagt agatgttctg ggatcatgaa gccactcacc ccagcagatt
     3721 ttaggtttac caaaaaactg gcctttgatt atcacggcaa tgagcttact ccgtcatccc
     3781 agtatgcatt taagttcaag gattattgtc ccgaggtttt ccgggagctt agagcactgt
     3841 ttggcctcga tcctgccgat tatctggtgt ctctcacatc taaatacatc ctctcagaac
     3901 tgaatagccc aggaaaatct gggtcatttt tttattatag tcgcgattac aagtatatta
     3961 ttaagactat tcaccactct gaacacatcc acctccggaa acacatccaa gaatactata
     4021 accatgtccg cgataacccg aatactctta tttgccaatt ttatggcctg catcgcgtca
     4081 agatgccaat ctccttccag aacaaaatca aacatcgaaa aatatatttc ctggtgatga
     4141 acaacctgtt ccccccacat cttgatattc acatcaccta tgatctcaag ggatccacat
     4201 ggggcaggtt caccaatctt gacaaagagc gcttggctaa agatcgcagc taccgcccag
     4261 tgatgaaaga ccttaactgg ctggaggagg gtcaaaagat taagttcggg ccactgaaga
     4321 aaaagacatt ccttacccag cttaagaagg atgtagagct tctggccaaa ctgaacacaa
     4381 tggattacag cctgcttatt ggcatccacg atatcaataa agccaaagag gacgacttgc
     4441 aattggcgga cactgctagc attgaagaac agcctcaaac gcagggccct attaggacag
     4501 gcacaggcac agttgtacgg catttcttcc gcgagttcga gggcggcata agagctagcg
     4561 accagtttaa caacgacgtc gaccttattt attatgtggg gataatcgac tttctgacaa
     4621 actacagtgt catgaaaaaa ctcgagactt tttggcgctc acttcgacac gacactaaat
     4681 tggtcagcgc tatacctcca cgggattatg ctaatcgctt ttacgagttt atcgaggatt
     4741 ccgtctaacg ggatccaccg gatctagata actgatcata atcagccata ccacatttgt
     4801 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     4861 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     4921 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     4981 caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg
     5041 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     5101 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     5161 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     5221 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     5281 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     5341 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     5401 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     5461 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5521 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5581 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     5641 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     5701 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     5761 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     5821 gcccagttcc gcccattctc cgccccatgg ctgactaatt t
//
