LOCUS       sequence-251760-        8404 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Inducible expression of Shark-mCherry in S2 cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8404)
  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 8404)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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 Shark-mCherry
FEATURES             Location/Qualifiers
     source          1..8404
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..40
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     234..256
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     248..265
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     249..265
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        271..697
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     CDS             732..749
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3657..4364
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     4067..4086
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(4384..4401)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4403..4537
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             5012..5611
                     /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(5012..5031)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5508..5528
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(5744..5760)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5745..5761)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    5776..5910
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6053..6069)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6053..6069)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6066..6088)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6077..6093
                     /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(6101..6107,6108..6125,6126..6131))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6146..6167
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6284..6301)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6455..7043)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6535..6554)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7214..8005,8006..8074))
                     /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     7837..7856
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8075..8179)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8247..8265
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(8303..8325)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg
       61 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt
      121 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
      181 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
      241 acgacgttgt aaaacgacgg ccagtgaatt aattcgttgc aggacaggat gtggtgcccg
      301 atgtgactag ctctttgctg caggccgtcc tatcctctgg ttccgataag agacccagaa
      361 ctccggcccc ccaccgccca ccgccacccc catacatatg tggtacgcaa gtaagagtgc
      421 ctgcgcatgc cccatgtgcc ccaccaagag ctttgcatcc catacaagtc cccaaagtgg
      481 agaaccgaac caattcttcg cgggcagaac aaaagcttct gcacacgtct ccactcgaat
      541 ttggagccgg ccggcgtgtg caaaagaggt gaatcgaacg aaagacccgt gtgtaaagcc
      601 gcgtttccaa aatgtataaa accgagagca tctggccaat gtgcatcagt tgtggtcagc
      661 agcaaaatca agtgaatcat ctcagtgcaa ctaaaggggg gatctagatc ggggtaccta
      721 ctagtgaaat gcaccatcac catcatcacg aagatcaagt tgacccaaga cttattgacg
      781 gaaaaggagg aggaggccgg cccatgagtc gcgatagtga tcccatgaaa tggtaccacg
      841 gcaacctgtc ccgcgaggca gccgacgaac tgctaaagca gggttacgag gacgggacat
      901 ttctggtgcg cgagagcagc acagcggccg gggactttgt gctgagcctg ctctgccagg
      961 gtgaggtgtg tcactaccag gtgcggcgac atggcggcga ggacgccttc ttctccatcg
     1021 acgacaaggt acagacgaag atcctgcacg gcctggacac gctcgtcgac tactatcagc
     1081 aggcggccaa cggtctgccc accaagctga cggtgccctt gatccgcgat ctgccgccgc
     1141 acaacactcg cagccacgga gtgaccaatc tgctgcatcg cgccacaagt aaaaacgaga
     1201 gcaaggtcgt cttcgagctg ctcaagtgcg gctatcggaa tttcgatgcc aagaatcagg
     1261 atggacagac ggctctgcat ctggctgcac tccactcgga cgaggatata ctgaaacacc
     1321 tgttgaatgc aaaggtccag gtcaacagct ccgactcctt tggctgccag ccattgcact
     1381 acgccgctcg ctccaaaccg gccagtttca tacgcacact catctcggca caggccaatg
     1441 tccagggcag gaacatcgac aacggctatg tgccgctgca tgaggctgcc aagcacggca
     1501 acctggaggc tgtgcaagag ctgctcctgg ctgaggctcc gccgttgccc agaacaagtt
     1561 cgggtgaatt tccattcgat ctggccaagg aagcgggcca aacggcagta gaagaattcc
     1621 tgctcaatta caagctgcca ccggcaaata ccacaagaga ccagtggtat catggcactc
     1681 tgaccagaga agaggcagta gccatactaa aaaagcatgc caaggagcta ttggccaagc
     1741 aacctgaagt agacacctcc ggctgctttc tggttcgcta ctccgagtcg ccggcggcct
     1801 cgggattggt tctaacattg cttagtgatc aggtggtcaa gaattttcgt atttcgcaag
     1861 ctgatctcta ccagaatggt aacaaggtgc agtcgggcgg gtccaagttt ctgtacattg
     1921 atgacggtcc ctactggcca agcgttgaac atctaatagc ccacttcatg cgtttctcgt
     1981 acgggttgcc agtgagtctc aagtatccgg ttccgccaca gcccaagcca gaggtgccat
     2041 cgtttgccac aatacctcga tctaatatga agccaaaagc agcatcgccc gctaccccgc
     2101 ccactccagt atcgccacat tcgcatcatc aacaccccca cgttcccgcc ctgaccataa
     2161 cgaagaaaaa gcagaaggag aacagtagct caatgttcaa cactctgcga ctgacgagtc
     2221 cgaagaaagc gctattcgac atgaacagcc tgcgcaaaaa caagtccaag ggcaaacgca
     2281 gtgactccga gagcagtgtg agtggttctt tggccagaac ggagcaggag ctgcaggcag
     2341 cggcacccat gctgaagagc ctgtcgttct cgacggagtt ctcaacgttt aacgcggatg
     2401 gcgtaactgg ttctggtgcc gccgctgctg gcgaagtgta caacgtgccc aggaacaaca
     2461 cgcccattga gattgacctg ccaccgattg cccaaaagac tgaagccgaa gttgagtatt
     2521 tcacgaaaag tgatgtggcc atagagcgcg agcgagcagg tcagtggatc ggcaacggct
     2581 accagcccac agtggacgtg ctctccctgt tggatcagca gatcaaggcg cccgccgtgg
     2641 cgcgactaaa ctcgcttggt ccgaacgcat caacggaaag tgagatggcc agctatcttc
     2701 atcgcaagtg cagcggtacg ccaagcacac caagcgccac tgaggtggag gccgccaagc
     2761 tgagattctt tattgagcca gaaaaactag tgctggatag agagatcggg cacggggaat
     2821 tcggttcggt gcacagtgga tggctgctga gaaagagcgg tgccggcgag gaatcgcgct
     2881 tggaggtggc cattaagatg cttagcgacg agcacagcaa caaacaggag ttcctgcgcg
     2941 aagcatcagt tatgatgcgg ctggagcaca agtgcattgt gcgattgatt ggcatttcga
     3001 agggtgaaat gctaatgatg gttcaagagc tagctccgct gggctccatg ctgcagtaca
     3061 tactggatca tggtcatgag atcacggcca atgcggagct gaaagtttgg gcgtcacaga
     3121 ttgcgtgtgg catgcattat ctggaatcac agcattttgt gcaccgcgat ctggccgcca
     3181 gaaatatatt gctgaccgca cgccaccagg cgaagatcag tgacttcggc atgtcgcgat
     3241 cactgagacc cggtagcacg gaataccaat tcacccaggg cggacgttgg cccatccgct
     3301 ggtacgcacc ggaaagcttc aacctgggca tcttctcgca cgccagtgat gtgtggtcct
     3361 ttggcgtgac catctgggag atgttttccc tcggcgcacc gccatatggg gaaatatcta
     3421 atgtcgatgc catcaagttg gtggacagtg gcgaacgcct gccgcagccc aatctctgtc
     3481 cagcttacat ctatgccgtg atgcagagct gctggaagga acgacccaaa gatcggccaa
     3541 cctttgtcta tctcaccgag tttttcgccc gcgatcctga ttaccaaaac ctgccggaat
     3601 tggtgcaaac ggttcacatt cggcgcgccg gcggccgctc gagtctagag ggccccgtga
     3661 gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca
     3721 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     3781 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct
     3841 gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg
     3901 acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
     3961 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt
     4021 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga
     4081 agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga
     4141 agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca
     4201 agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca
     4261 agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg
     4321 agggccgcca ctccaccggc ggcatggacg agctgtacaa gtaagtttaa acccgctgat
     4381 cagcctcgac tgtgccttct aagatccaga catgataaga tacattgatg agtttggaca
     4441 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     4501 tttatttgta accattataa gctgcaataa acaagttaac tctagaggat ccggtgcgtg
     4561 gtggtttcat gcttctggga acggcaaatg ggtttaggat tgggaacccc tcatcatctg
     4621 ttggaatata ctattcaacc tacaaaagta acgttaaaca acactacttt atatttgata
     4681 tgaatggcca caccttttat gccataaaac atattgtaag agaataccac tctttttatt
     4741 ccttctttcc ttcttgtacg ttttttgctg taagtaggtc gtggtgctgg tgttgcagtt
     4801 gaaataactt aaaatataaa tcataaaact caaacataaa cttgactatt tatttattta
     4861 ttaagaaagg aaatataaat tataaattac aacagatccc ccgggctgca ggaattcgat
     4921 agcttgcatg cctgcaggtc ggccgccacg accggtgccg ccaccatccc ctgacccacg
     4981 cccctgaccc ctcacaagga gacgaccttc catgaccgag tacaagccca cggtgcgcct
     5041 cgccacccgc gacgacgtcc cccgggccgt acgcaccctc gccgccgcgt tcgccgacta
     5101 ccccgccacg cgccacaccg tcgacccgga ccgccacatc gagcgggtca ccgagctgca
     5161 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     5221 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     5281 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     5341 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     5401 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     5461 ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt
     5521 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     5581 ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc cacgacccgc agcgcccgac
     5641 cgaaaggagc gcacgacccc atggctccga ccgaagccac ccggggcggc cccgccgacc
     5701 ccgcacccgc ccccgaggcc caccgactct agatcaagct tatcgatacc gtcgacctcg
     5761 aggccagcta gcttgaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5821 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5881 ctcatcaatg tatcttatca tgtctggatc gggtacccaa ttgcattcat tttatgtttc
     5941 aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta
     6001 tggctgatta tgatcagtcg acctgcaggc atgcaagctt ggcgtaatca tggtcatagc
     6061 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     6121 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     6181 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     6241 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     6301 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     6361 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     6421 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     6481 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     6541 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     6601 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     6661 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     6721 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     6781 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     6841 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     6901 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     6961 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     7021 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     7081 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     7141 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     7201 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     7261 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     7321 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     7381 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     7441 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     7501 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     7561 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     7621 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     7681 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     7741 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     7801 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     7861 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     7921 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     7981 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     8041 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     8101 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     8161 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     8221 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc
     8281 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     8341 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     8401 ggtg
//
