LOCUS       sequence-251731-        5535 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of Lifeact-TagBFP in S2 cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5535)
  AUTHORS   Williamson AP, Vale RD
  TITLE     Spatial control of Draper receptor signaling initiates apoptotic 
            cell engulfment.
  JOURNAL   J Cell Biol. 2018 Nov 5;217(11):3977-3992. doi: 
            10.1083/jcb.201711175. Epub 2018 Aug 23.
  PUBMED    30139739
REFERENCE   2  (bases 1 to 5535)
  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: 
            "10.1083/jcb.201711175"; journalName: "J Cell Biol"; date: 
            "2018-11-5- 5"; volume: "217"; issue: "11"; pages: "3977-3992"
COMMENT     Sequence Label: pMT Lifeact-TagBFP
FEATURES             Location/Qualifiers
     source          1..5535
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     38..60
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     52..69
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     53..69
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        75..501
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     CDS             533..583
                     /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"
     CDS             596..1297
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     primer_bind     complement(1319..1336)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1338..1472
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             1947..2546
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1947..1966)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2443..2463
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(2679..2695)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2680..2696)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2711..2845
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2988..3004)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2988..3004)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3001..3023)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3012..3028
                     /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(3036..3042,3043..3060,3061..3066))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3081..3102
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3219..3236)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3390..3978)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3470..3489)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4149..4940,4941..5009))
                     /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     4772..4791
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5010..5114)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5182..5200
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5238..5260)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5360..5379
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
       61 acggccagtg aattaattcg ttgcaggaca ggatgtggtg cccgatgtga ctagctcttt
      121 gctgcaggcc gtcctatcct ctggttccga taagagaccc agaactccgg ccccccaccg
      181 cccaccgcca cccccataca tatgtggtac gcaagtaaga gtgcctgcgc atgccccatg
      241 tgccccacca agagctttgc atcccataca agtccccaaa gtggagaacc gaaccaattc
      301 ttcgcgggca gaacaaaagc ttctgcacac gtctccactc gaatttggag ccggccggcg
      361 tgtgcaaaag aggtgaatcg aacgaaagac ccgtgtgtaa agccgcgttt ccaaaatgta
      421 taaaaccgag agcatctggc caatgtgcat cagttgtggt cagcagcaaa atcaagtgaa
      481 tcatctcagt gcaactaaag gggggatcta gatcggggta cctactagtc aaatgggtgt
      541 cgcagatttg atcaagaaat tcgaaagcat ctcaaaggaa gaaggttctg gttctatgag
      601 cgagctgatt aaggagaaca tgcacatgaa gctgtacatg gagggcaccg tggacaacca
      661 tcacttcaag tgcacatccg agggcgaagg caagccctac gagggcaccc agaccatgag
      721 aatcaaggtg gtcgagggcg gccctctccc cttcgccttc gacatcctgg ctactagctt
      781 cctctacggc agcaagacct tcatcaacca cacccagggc atccccgact tcttcaagca
      841 gtccttccct gagggcttca catgggagag agtcaccaca tacgaagacg ggggcgtgct
      901 gaccgctacc caggacacca gcctccagga cggctgcctc atctacaacg tcaagatcag
      961 aggggtgaac ttcacatcca acggccctgt gatgcagaag aaaacactcg gctgggaggc
     1021 cttcaccgag acgctgtacc ccgctgacgg cggcctggaa ggcagaaacg acatggccct
     1081 gaagctcgtg ggcgggagcc atctgatcgc aaacatcaag accacatata gatccaagaa
     1141 acccgctaag aacctcaaga tgcctggcgt ctactatgtg gactacagac tggaaagaat
     1201 caaggaggcc aacaacgaga cctacgtcga gcagcacgag gtggcagtgg ccagatactg
     1261 cgacctccct agcaaactgg ggcacaagct taattaagtt taaacgcccg ctgatcagcc
     1321 tcgactgtgc cttctaagat ccagacatga taagatacat tgatgagttt ggacaaacca
     1381 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     1441 ttgtaaccat tataagctgc aataaacaag ttaactctag aggatccggt gcgtggtggt
     1501 ttcatgcttc tgggaacggc aaatgggttt aggattggga acccctcatc atctgttgga
     1561 atatactatt caacctacaa aagtaacgtt aaacaacact actttatatt tgatatgaat
     1621 ggccacacct tttatgccat aaaacatatt gtaagagaat accactcttt ttattccttc
     1681 tttccttctt gtacgttttt tgctgtaagt aggtcgtggt gctggtgttg cagttgaaat
     1741 aacttaaaat ataaatcata aaactcaaac ataaacttga ctatttattt atttattaag
     1801 aaaggaaata taaattataa attacaacag atcccccggg ctgcaggaat tcgatagctt
     1861 gcatgcctgc aggtcggccg ccacgaccgg tgccgccacc atcccctgac ccacgcccct
     1921 gacccctcac aaggagacga ccttccatga ccgagtacaa gcccacggtg cgcctcgcca
     1981 cccgcgacga cgtcccccgg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg
     2041 ccacgcgcca caccgtcgac ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac
     2101 tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg
     2161 cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg
     2221 gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc
     2281 tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc
     2341 ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg
     2401 agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc cccttctacg
     2461 agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt
     2521 gcatgacccg caagcccggt gcctgacgcc cgccccacga cccgcagcgc ccgaccgaaa
     2581 ggagcgcacg accccatggc tccgaccgaa gccacccggg gcggccccgc cgaccccgca
     2641 cccgcccccg aggcccaccg actctagatc aagcttatcg ataccgtcga cctcgaggcc
     2701 agctagcttg aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     2761 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     2821 caatgtatct tatcatgtct ggatcgggta cccaattgca ttcattttat gtttcaggtt
     2881 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
     2941 gattatgatc agtcgacctg caggcatgca agcttggcgt aatcatggtc atagctgttt
     3001 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     3061 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     3121 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     3181 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3241 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3301 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     3361 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3421 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     3481 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     3541 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     3601 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     3661 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     3721 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     3781 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     3841 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     3901 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     3961 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4021 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4081 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4141 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4201 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4261 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     4321 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4381 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4441 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     4501 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     4561 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     4621 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     4681 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     4741 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     4801 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     4861 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     4921 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     4981 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5041 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     5101 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     5161 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc
     5221 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     5281 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     5341 cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg
     5401 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgcca ttcgccattc
     5461 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
     5521 gcgaaagggg gatgt
//
