LOCUS       sequence-251722-        6136 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of vCrk truncation in S2 cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6136)
  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 6136)
  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 vCrk-GFP
FEATURES             Location/Qualifiers
     source          1..6136
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     112..134
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     126..143
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     127..143
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        149..575
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     CDS             610..627
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             628..663
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             1261..1974
                     /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             1261..1971
                     /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(1288..1316)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1911..1931
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(1994..2011)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2013..2147
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             2622..3221
                     /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(2622..2641)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3118..3138
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(3354..3370)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3355..3371)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    3386..3520
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3663..3679)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3663..3679)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3676..3698)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3687..3703
                     /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(3711..3717,3718..3735,3736..3741))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3756..3777
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3894..3911)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4065..4653)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4145..4164)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4824..5615,5616..5684))
                     /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     5447..5466
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5685..5789)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5857..5875
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5913..5935)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6035..6054
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg
       61 cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac
      121 gacgttgtaa aacgacggcc agtgaattaa ttcgttgcag gacaggatgt ggtgcccgat
      181 gtgactagct ctttgctgca ggccgtccta tcctctggtt ccgataagag acccagaact
      241 ccggcccccc accgcccacc gccaccccca tacatatgtg gtacgcaagt aagagtgcct
      301 gcgcatgccc catgtgcccc accaagagct ttgcatccca tacaagtccc caaagtggag
      361 aaccgaacca attcttcgcg ggcagaacaa aagcttctgc acacgtctcc actcgaattt
      421 ggagccggcc ggcgtgtgca aaagaggtga atcgaacgaa agacccgtgt gtaaagccgc
      481 gtttccaaaa tgtataaaac cgagagcatc tggccaatgt gcatcagttg tggtcagcag
      541 caaaatcaag tgaatcatct cagtgcaact aaagggggga tctagatcgg ggtacctact
      601 agtgaaatgc accatcacca tcatcacgaa gatcaagttg acccaagact tattgacgga
      661 aaaggaggag gaggccggcc catggataca tttgacgttt ctgataggaa cagctggtac
      721 tttggtccca tgtctagaca ggatgctact gaagttttga tgaacgaacg cgagcgggga
      781 gtgtttttag tccgtgatag taactcgata gcaggggatt atgtactttg tgtaagagaa
      841 gatacaaaag ttagcaacta catcattaac aaagttcaac aacaggatca aatcgtttac
      901 cgcattgggg atcagtcttt tgacaatcta ccgaaactct taacttttta cactcttcat
      961 tatttggata caaccccttt aaaacggcct gcgtgtagaa gggtggaaaa agtaatagga
     1021 aagttcgatt tcgttggcag cgatcaagat gatttacctt ttcaaagagg tgaagtttta
     1081 acaatagttc gaaaagacga ggatcaatgg tggactgcgc gtaactcctc ggggaaaatt
     1141 ggtcaaatac cggttcccta tatacaacag tatgacgatt atatggatga agatgctatt
     1201 gataaaaacg aaccttccat ttcggggcgc gccggcggcc gctcgagtct agagggcccc
     1261 agcaaaggag aagaactttt cactggagtt gtcccaattc ttgttgaatt agatggtgat
     1321 gttaatgggc acaaattttc tgtccgtgga gagggtgaag gtgatgctac aaacggaaaa
     1381 ctcaccctta aatttatttg cactactgga aaactacctg ttccgtggcc aacacttgtc
     1441 actactctga cctatggtgt tcaatgcttt tcccgttatc cggatcacat gaaacggcat
     1501 gactttttca agagtgccat gcccgaaggt tatgtacagg aacgcactat atctttcaaa
     1561 gatgacggga cctacaagac gcgtgctgaa gtcaagtttg aaggtgatac ccttgttaat
     1621 cgtatcgagt taaagggtat tgattttaaa gaagatggaa acattcttgg acacaaactc
     1681 gagtacaact ttaactcaca caatgtatac atcacggcag acaaacaaaa gaatggaatc
     1741 aaagctaact tcaaaattcg ccacaacgtt gaagatggtt ccgttcaact agcagaccat
     1801 tatcaacaaa atactccaat tggcgatggc cctgtccttt taccagacaa ccattacctg
     1861 tcgacacaat ctgtcctttc gaaagatccc aacgaaaagc gtgaccacat ggtccttctt
     1921 gagtttgtaa ctgctgctgg gattacacat ggcatggatg agctctacaa ataggtttaa
     1981 acccgctgat cagcctcgac tgtgccttct aagatccaga catgataaga tacattgatg
     2041 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     2101 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac tctagaggat
     2161 ccggtgcgtg gtggtttcat gcttctggga acggcaaatg ggtttaggat tgggaacccc
     2221 tcatcatctg ttggaatata ctattcaacc tacaaaagta acgttaaaca acactacttt
     2281 atatttgata tgaatggcca caccttttat gccataaaac atattgtaag agaataccac
     2341 tctttttatt ccttctttcc ttcttgtacg ttttttgctg taagtaggtc gtggtgctgg
     2401 tgttgcagtt gaaataactt aaaatataaa tcataaaact caaacataaa cttgactatt
     2461 tatttattta ttaagaaagg aaatataaat tataaattac aacagatccc ccgggctgca
     2521 ggaattcgat agcttgcatg cctgcaggtc ggccgccacg accggtgccg ccaccatccc
     2581 ctgacccacg cccctgaccc ctcacaagga gacgaccttc catgaccgag tacaagccca
     2641 cggtgcgcct cgccacccgc gacgacgtcc cccgggccgt acgcaccctc gccgccgcgt
     2701 tcgccgacta ccccgccacg cgccacaccg tcgacccgga ccgccacatc gagcgggtca
     2761 ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg
     2821 cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg
     2881 tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc
     2941 aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca
     3001 ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg
     3061 gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca
     3121 acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag
     3181 gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc cacgacccgc
     3241 agcgcccgac cgaaaggagc gcacgacccc atggctccga ccgaagccac ccggggcggc
     3301 cccgccgacc ccgcacccgc ccccgaggcc caccgactct agatcaagct tatcgatacc
     3361 gtcgacctcg aggccagcta gcttgaactt gtttattgca gcttataatg gttacaaata
     3421 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     3481 tttgtccaaa ctcatcaatg tatcttatca tgtctggatc gggtacccaa ttgcattcat
     3541 tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac
     3601 aaatgtggta tggctgatta tgatcagtcg acctgcaggc atgcaagctt ggcgtaatca
     3661 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3721 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3781 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3841 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3901 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     3961 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4021 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4081 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4141 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4201 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4261 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4321 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4381 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4441 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4501 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4561 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4621 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4681 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4741 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4801 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4861 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4921 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4981 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5041 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5101 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5161 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5221 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5281 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5341 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5401 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5461 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5521 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5581 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5641 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5701 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5761 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     5821 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     5881 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     5941 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     6001 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
     6061 gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     6121 cgccattcgc cattca
//
