LOCUS       sequence-446761-        5463 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses LifeAct-mScarlet-scramble to serve as control plasmid for 
            LifeAct-MTED.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5463)
  AUTHORS   Holland ED, Miller HL, Millette MM, Taylor RJ, Drucker GL, Dent EW
  TITLE     A methodology for specific disruption of microtubule polymerization 
            into dendritic spines.
  JOURNAL   Mol Biol Cell. 2024 Jun 1;35(6):mr3. doi: 10.1091/mbc.E24-02-0093. 
            Epub 2024 Apr 17.
  PUBMED    38630519
REFERENCE   2  (bases 1 to 5463)
  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: "Mol Biol 
            Cell."; date: "2024-06-1"; volume: "35(6):mr3. doi"; pages: " 
            10.1091/mbc.E24-02-0093. Epub 2024 Apr 17"
COMMENT     Sequence Label: LifeAct-scramble
FEATURES             Location/Qualifiers
     source          1..5463
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    121..255
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(158..177)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     212..231
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(323..340)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(494..1082)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(574..593)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1253..2044,2045..2113))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1876..1895
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2114..2218)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2274..2588
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    2282..2300
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2318..2336
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2353..2371
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2389..2407
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2425..2443
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2460..2478
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2496..2514
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2561..2585
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2614..2623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2620..2670
                     /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"
     promoter        3480..3990
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3909..3927
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             join(4000..4617,4618..4623,4624..4659,4660..4662,4663..4698,
                     4699..4701,4702..4737,4738..4746)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     polyA_signal    4794..4849
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4848..4867)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5210..5226)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5210..5226)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5223..5245)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5234..5250
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5258..5264,5265..5282,5283..5288))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5303..5324
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5383..5463,1..115)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      join(5429..5463,1..101)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
       61 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
      121 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
      181 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
      241 tatcatgtct ggatccgctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
      301 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
      361 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      421 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      481 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      541 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      601 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      661 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      721 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
      781 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      841 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      901 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
      961 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     1021 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     1081 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     1141 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     1201 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     1261 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1321 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     1381 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     1441 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1501 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1561 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1621 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1681 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1741 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1801 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1861 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1921 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1981 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     2041 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     2101 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     2161 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     2221 catttccccg aaaagtgcca cctggtcgtc gactggccgc gaatcgatat gtcgagttta
     2281 ctccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga tagagaacga
     2341 tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttactc cctatcagtg
     2401 atagagaacg tatgtcgagt ttactcccta tcagtgatag agaacgtatg tcgagtttat
     2461 ccctatcagt gatagagaac gtatgtcgag tttactccct atcagtgata gagaacgtat
     2521 gtcgaggtag gcgtgtacgg tgggaggcct atataagcag agctcgttta gtgaaccgtc
     2581 agatcgcctg gagaaaccgg ttatactgcg gccgccacca tgggcgtggc ggatctgatt
     2641 aagaagttcg aaagcattag caaagaagaa ggctctggtt ccggcactag tgtgagtaaa
     2701 ggagaggccg ttataaaaga gttcatgcgg ttcaaggtac acatggaggg atcaatgaac
     2761 ggacacgagt tcgaaatcga aggggaagga gagggcaggc cctatgaggg tactcagacc
     2821 gccaaactca aagtgacaaa gggaggccct ttgcctttct cttgggacat cttgtcacca
     2881 caatttatgt atggctcccg agcatttata aaacacccag cagacatacc tgattactac
     2941 aagcagtctt tcccagaagg ttttaaatgg gaaagggtca tgaactttga ggatggtggt
     3001 gctgtgacgg tgacgcagga tacttctttg gaggacggaa cactcatcta caaggtaaag
     3061 ctcaggggca cgaacttccc gccggacgga ccagtcatgc agaaaaagac tatgggatgg
     3121 gaggcttcca ccgaacggtt gtatcccgag gacggtgtct tgaagggaga tataaaaatg
     3181 gctcttcggc tgaaagatgg aggcagatac ttggcagatt ttaagaccac gtataaggca
     3241 aaaaaaccag tgcaaatgcc gggagcatat aacgttgacc gcaaactgga catcacctca
     3301 cataacgagg attatacagt ggtagaacag tacgaaagaa gcgaagggag gcacagcacc
     3361 gggggtatgg atgagctgta taagggcagc ggcagcggca gcatgattac cgcgccgcgc
     3421 gaatttgatt atctgaacct gcgcgcgggc ctgagcatga cctgaacgcg tcatgctccg
     3481 gggttggggt tgcgcctttt ccaaggcagc cctgggtttg cgcagggacg cggctgctct
     3541 gggcgtggtt ccgggaaacg cagcggcgcc gaccctgggt ctcgcacatt cttcacgtcc
     3601 gttcgcagcg tcacccggat cttcgccgct acccttgtgg gccccccggc gacgcttcct
     3661 gctccgcccc taagtcggga aggttccttg cggttcgcgg cgtgccggac gtgacaaacg
     3721 gaagccgcac gtctcactag taccctcgca gacggacagc gccagggagc aatggcagcg
     3781 cgccgaccgc gatgggctgt ggccaatagc ggctgctcag cagggcgcgc cgagagcagc
     3841 ggccgggaag gggcggtgcg ggaggcgggg tgtggggcgg tagtgtgggc cctgttcctg
     3901 cccgcgcggt gttccgcatt ctgcaagcct ccggagcgca cgtcggcagt cggctccctc
     3961 gttgaccgaa tcaccgacct ctctccccag caattcacca tgtctagact ggacaagagc
     4021 aaagtcataa acggagctct ggaattactc aatggtgtcg gtatcgaagg cctgacgaca
     4081 aggaaactcg ctcaaaagct gggagttgag cagcctaccc tgtactggca cgtgaagaac
     4141 aagcgggccc tgctcgatgc cctgccaatc gagatgctgg acaggcatca tacccacttc
     4201 tgccccctgg aaggcgagtc atggcaagac tttctgcgga acaacgccaa gtcataccgc
     4261 tgtgctctcc tctcacatcg cgacggggct aaagtgcatc tcggcacccg cccaacagag
     4321 aaacagtacg aaaccctgga aaatcagctc gcgttcctgt gtcagcaagg cttctccctg
     4381 gagaacgcac tgtacgctct gtccgccgtg ggccacttta cactgggctg cgtattggag
     4441 gaacaggagc atcaagtagc aaaagaggaa agagagacac ctaccaccga ttctatgccc
     4501 ccacttctga gacaagcaat tgagctgttc gaccggcagg gagccgaacc tgccttcctt
     4561 ttcggcctgg aactaatcat atgtggcctg gagaaacagc taaagtgcga aagcggcggg
     4621 ccgaccgacg cccttgacga ttttgactta gacatgctcc cagccgatgc ccttgacgac
     4681 tttgaccttg atatgctgcc tgctgacgct cttgacgatt ttgaccttga catgctcccc
     4741 gggtaactaa gtaaggatcg atccaaggta ccttgagcat ctgacttctg gctaataaag
     4801 gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac
     4861 atatgggagg gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac
     4921 atatgcccat atgctggctg ccatgaacaa aggttggcta taaagaggtc atcagtatat
     4981 gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat
     5041 tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca
     5101 tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct
     5161 tctcttatgg agatccctcg acctgcagcc caagcttggc gtaatcatgg tcatagctgt
     5221 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     5281 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     5341 tgcccgcttt ccagtcggga aacctgtcgt gccagcggat ccgcatctca attagtcagc
     5401 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     5461 ttc
//
