LOCUS       sequence-177520-        4771 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Localization by Fluorescence microscopy of lifeAct fused to super 
            folder GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4771)
  TITLE     Unpublished Henriques Super-Resolution Microscopy Plasmids
REFERENCE   2  (bases 1 to 4771)
  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"
COMMENT     Sequence Label: RHPMP192 (pLifeAct sfGFP)
FEATURES             Location/Qualifiers
     source          1..4771
                     /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"
     CDS             639..689
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     regulatory      705..714
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(717..719,720..722,723..1436)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(762..783)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1370..1391
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1559..1680
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1596..1615)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1650..1669
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1687..2142)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1824..1845)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2036..2055
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2169..2273
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2275..2632
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2275..2466
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2298..2318)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2483..2618
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2667..3461
                     /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             2667..3461
                     /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(2721..2740)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3331..3350
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3649..3668)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3693..3740
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4069..4657
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4558..4577
                     /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 caagcttcga attccaccat gggcgtggcc gacctgatca
      661 agaagttcga gagcatcagc aaggaggagg gggatccacc ggtcgccacc atggtgatgg
      721 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
      781 acgtaaacgg ccacaagttc agcgtgcgcg gcgagggcga gggcgatgcc accaacggca
      841 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
      901 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcgcc
      961 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcagcttca
     1021 aggacgacgg cacctacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     1081 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     1141 tggagtacaa cttcaacagc cacaacgtct atatcaccgc cgacaagcag aagaacggca
     1201 tcaaggccaa cttcaagatc cgccacaacg tggaggacgg cagcgtgcag ctcgccgacc
     1261 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     1321 tgagcaccca gtccgtgctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     1381 tggagttcgt gaccgccgcc gggatcactc acggcatgga cgagctgtac aagtaaagcg
     1441 gccgcgactc tagatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     1501 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     1561 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     1621 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     1681 aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     1741 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     1801 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     1861 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     1921 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     1981 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     2041 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     2101 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     2161 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2221 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg
     2281 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     2341 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     2401 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     2461 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     2521 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     2581 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc
     2641 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     2701 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     2761 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     2821 acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca
     2881 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     2941 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     3001 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     3061 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     3121 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     3181 ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     3241 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     3301 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     3361 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     3421 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     3481 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     3541 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     3601 cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa
     3661 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     3721 cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     3781 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     3841 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     3901 catagcctca ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3961 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     4021 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     4081 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     4141 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     4201 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     4261 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     4321 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4381 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     4441 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     4501 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     4561 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4621 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4681 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4741 ttctgtggat aaccgtatta ccgccatgca t
//
