LOCUS       sequence-251717-        7081 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of p85-GFP in S2 cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7081)
  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 7081)
  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 p85-GFP
FEATURES             Location/Qualifiers
     source          1..7081
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     180..202
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     194..211
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     195..211
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        217..643
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     CDS             678..695
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             696..731
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2274..2987
                     /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="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             2274..2984
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(2301..2329)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2924..2944
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(3007..3024)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3026..3160
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             3635..4234
                     /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(3635..3654)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4131..4151
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4367..4383)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4368..4384)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    4399..4533
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4676..4692)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4676..4692)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4689..4711)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4700..4716
                     /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(4724..4730,4731..4748,4749..4754))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4769..4790
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4907..4924)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5078..5666)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5158..5177)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5837..6628,6629..6697))
                     /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     6460..6479
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6698..6802)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6870..6888
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6926..6948)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7048..7067
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
       61 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
      121 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
      181 ccagtcacga cgttgtaaaa cgacggccag tgaattaatt cgttgcagga caggatgtgg
      241 tgcccgatgt gactagctct ttgctgcagg ccgtcctatc ctctggttcc gataagagac
      301 ccagaactcc ggccccccac cgcccaccgc cacccccata catatgtggt acgcaagtaa
      361 gagtgcctgc gcatgcccca tgtgccccac caagagcttt gcatcccata caagtcccca
      421 aagtggagaa ccgaaccaat tcttcgcggg cagaacaaaa gcttctgcac acgtctccac
      481 tcgaatttgg agccggccgg cgtgtgcaaa agaggtgaat cgaacgaaag acccgtgtgt
      541 aaagccgcgt ttccaaaatg tataaaaccg agagcatctg gccaatgtgc atcagttgtg
      601 gtcagcagca aaatcaagtg aatcatctca gtgcaactaa aggggggatc tagatcgggg
      661 tacctactag tgaaatgcac catcaccatc atcacgaaga tcaagttgac ccaagactta
      721 ttgacggaaa aggaggagga ggccggccca tgcggccaca ggcgcccgga tcgctggtcg
      781 atcccaacga ggacgagcta cgcatggccc catggtactg gggccggata tcgagggagg
      841 aagccaagag catcctgcac ggaaagccgg acggcagctt tttggtgcgg gacgccctct
      901 cgatgaaggg cgagtacact ctgaccctga tgaaggacgg atgcgagaag ctaataaaga
      961 tatgccacat ggatcgcaag tacggcttta tcgagacgga cctgttcaat tcggtggtgg
     1021 agatgatcaa ctactacaag gagaactctc tgagcatgta caacaagacg ctggacataa
     1081 cactgagtaa tccgattgtg cgagcccgcg aggatgagga gtcgcagccg cacggggatc
     1141 tttgtctgct gagcaacgaa ttcatcagga cctgccagct gcttcagaat cttgagcaaa
     1201 atctcgagaa taagcgcaac tcattcaatg cgattcgaga ggagctgcag gagaagaagc
     1261 tgcaccagag cgtattcggg aatacagaga agatattccg caatcaaatc aagctcaatg
     1321 aatccttcat gaaggcgcca gcggatgccc cgtccacgga ggcaggagga gctggagatg
     1381 gagctaacgc agccgcatcc gcagcggcca atgccaacgc tcgccggagt ctgcaggagc
     1441 acaagcagac actcctcaat ctgctggatg ctcttcaggc caagggtcaa gtccttaacc
     1501 actacatgga gaacaagaag aaggaggagc tactgctcga gcgacagatc aacgccctga
     1561 agccggaatt acagatactg cagttgcgca aggacaagta cattgagcgc ctaaaaggat
     1621 tcaacctcaa ggacgacgat ctgaagatga tattgcagat gggcttcgac aagtggcagc
     1681 aactttatga gacagtctcc aaccagccgc acagcaacga agctctctgg ttgctaaagg
     1741 atgccaagag aaggaatgcg gaggagatgc ttaagggggc gccatcgggc accttcctga
     1801 ttcgagcgag agatgcgggt cactatgctc tgtccattgc ctgcaagaac atcgtgcagc
     1861 actgccttat ttacgagacg agcaccggct ttggatttgc tgccccttac aatatatatg
     1921 ccacgctgaa gagtctggtg gagcactatg ccaacaattc gctggaggag cataacgaca
     1981 cgctgaccac gacattgcgc tggcctgtct tgtactggaa gaacaacccg ctgcaagttc
     2041 agatgatcca attgcaggag gaaatggatc tggagtacga gcaggcagcc acgttgaggc
     2101 cgccgccaat gatgggcagc agtgcgccga ttcccacgag tcggtcccgt gaacatgacg
     2161 tcgtagatgg aactggaagc ctcgaggcgg aggcggctcc ggcctccatt tcgccctcca
     2221 acttcagcac atcgcagggg cgcgccggcg gccgctcgag tctagagggc cccagcaaag
     2281 gagaagaact tttcactgga gttgtcccaa ttcttgttga attagatggt gatgttaatg
     2341 ggcacaaatt ttctgtccgt ggagagggtg aaggtgatgc tacaaacgga aaactcaccc
     2401 ttaaatttat ttgcactact ggaaaactac ctgttccgtg gccaacactt gtcactactc
     2461 tgacctatgg tgttcaatgc ttttcccgtt atccggatca catgaaacgg catgactttt
     2521 tcaagagtgc catgcccgaa ggttatgtac aggaacgcac tatatctttc aaagatgacg
     2581 ggacctacaa gacgcgtgct gaagtcaagt ttgaaggtga tacccttgtt aatcgtatcg
     2641 agttaaaggg tattgatttt aaagaagatg gaaacattct tggacacaaa ctcgagtaca
     2701 actttaactc acacaatgta tacatcacgg cagacaaaca aaagaatgga atcaaagcta
     2761 acttcaaaat tcgccacaac gttgaagatg gttccgttca actagcagac cattatcaac
     2821 aaaatactcc aattggcgat ggccctgtcc ttttaccaga caaccattac ctgtcgacac
     2881 aatctgtcct ttcgaaagat cccaacgaaa agcgtgacca catggtcctt cttgagtttg
     2941 taactgctgc tgggattaca catggcatgg atgagctcta caaataggtt taaacccgct
     3001 gatcagcctc gactgtgcct tctaagatcc agacatgata agatacattg atgagtttgg
     3061 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
     3121 tgctttattt gtaaccatta taagctgcaa taaacaagtt aactctagag gatccggtgc
     3181 gtggtggttt catgcttctg ggaacggcaa atgggtttag gattgggaac ccctcatcat
     3241 ctgttggaat atactattca acctacaaaa gtaacgttaa acaacactac tttatatttg
     3301 atatgaatgg ccacaccttt tatgccataa aacatattgt aagagaatac cactcttttt
     3361 attccttctt tccttcttgt acgttttttg ctgtaagtag gtcgtggtgc tggtgttgca
     3421 gttgaaataa cttaaaatat aaatcataaa actcaaacat aaacttgact atttatttat
     3481 ttattaagaa aggaaatata aattataaat tacaacagat cccccgggct gcaggaattc
     3541 gatagcttgc atgcctgcag gtcggccgcc acgaccggtg ccgccaccat cccctgaccc
     3601 acgcccctga cccctcacaa ggagacgacc ttccatgacc gagtacaagc ccacggtgcg
     3661 cctcgccacc cgcgacgacg tcccccgggc cgtacgcacc ctcgccgccg cgttcgccga
     3721 ctaccccgcc acgcgccaca ccgtcgaccc ggaccgccac atcgagcggg tcaccgagct
     3781 gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga
     3841 cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc
     3901 cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat
     3961 ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg
     4021 cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga
     4081 ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc
     4141 cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg
     4201 cacctggtgc atgacccgca agcccggtgc ctgacgcccg ccccacgacc cgcagcgccc
     4261 gaccgaaagg agcgcacgac cccatggctc cgaccgaagc cacccggggc ggccccgccg
     4321 accccgcacc cgcccccgag gcccaccgac tctagatcaa gcttatcgat accgtcgacc
     4381 tcgaggccag ctagcttgaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     4441 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     4501 aaactcatca atgtatctta tcatgtctgg atcgggtacc caattgcatt cattttatgt
     4561 ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
     4621 gtatggctga ttatgatcag tcgacctgca ggcatgcaag cttggcgtaa tcatggtcat
     4681 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4741 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4801 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4861 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4921 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4981 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5041 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5101 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5161 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5221 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5281 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5341 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5401 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5461 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5521 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5581 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5641 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5701 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5761 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5821 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5881 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5941 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6001 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6061 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6121 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6181 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6241 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6301 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6361 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6421 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6481 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6541 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6601 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6661 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6721 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6781 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     6841 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg
     6901 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     6961 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     7021 gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     7081 a
//
