LOCUS       sequence-251773-        6007 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of His10-Draper delta ITAM mutant for purification from 
            E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6007)
  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 6007)
  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: pET His10-Draper delta ITAM
FEATURES             Location/Qualifiers
     source          1..6007
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     37..56
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     140..159
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        140..158
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    159..183
                     /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)."
     RBS             198..220
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             239..268
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             278..295
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             299..331
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             1011..1028
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1081..1099)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1095..1142
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1179..1634
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1266..1285)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1476..1497
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1726..2541)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1726..2541)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2449..2468
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2533..2552
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2663..3251
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3152..3171
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3405..3422
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3437..3576
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3502..3521)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3662..3684
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3678..3869)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4678..5760)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5722..5741
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5761..5838)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc
       61 agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga
      121 gatctcgatc ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat
      181 tcccctctag aaataatttt gtttaacttt aagaaggaga tataccatgg gcagcagcca
      241 tcatcatcat catcatcatc accatcacag cagcggcctg gtgccgcgcg gcagccatat
      301 ggctagcatg actggtggac agcaaatggg tcgcggatcc cgccgacgtg tgtccaatct
      361 aaaaaccgaa atcgcccatg tacactacac ccatgatacg aatcccccga gctggccgcc
      421 caaccacaac ttcgacaatc ccgtgtacgg catgcaggcg gaaacgcgtc tcctgcccaa
      481 caatatgcgt tccaagatga acaactttga ccagaggagc acgatgagca cggactacgg
      541 cgatgactgc aatgccagtg gcagggtggg tagctattcg atcaactaca accacgacct
      601 gctaacgaag aacttgaacg cggaccggac caatccgatt gtgtacaacg aatccctcaa
      661 agaggagcat gtcttcgacg agatcaagca caaggagggc tacaaggatc ccgatgaatt
      721 cgatcacctg gactactcac gacccagcac ctcgcagaag ccgcactacc accgcatgaa
      781 cgacgccatg ctcaacatca accaggacga ggagaagccc agcaacgtga agaacatgac
      841 tgtgctgctg aacaaaccgc tgccgcccac ggaaccggag ccacaacacg agtgcttcga
      901 caacacgaac accaacctgg acaacgtgtc gacggcgtcg cctagttcca gtccgaaatt
      961 tctaaagtaa gaattcgagc tccgtcgaca agcttgcggc cgcactcgag caccaccacc
     1021 accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     1081 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     1141 tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta gcggcgcatt
     1201 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     1261 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     1321 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     1381 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     1441 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     1501 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     1561 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     1621 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     1681 tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga aaaactcatc
     1741 gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat atttttgaaa
     1801 aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga tggcaagatc
     1861 ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta atttcccctc
     1921 gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat ccggtgagaa
     1981 tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat tacgctcgtc
     2041 atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct gagcgagacg
     2101 aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca accggcgcag
     2161 gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt ctaatacctg
     2221 gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag gagtacggat
     2281 aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc tgaccatctc
     2341 atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact ctggcgcatc
     2401 gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat cgcgagccca
     2461 tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag agcaagacgt
     2521 ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag cagacagttt
     2581 tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2641 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2701 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2761 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     2821 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2881 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     2941 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3001 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3061 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3121 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3181 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3241 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3301 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3361 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3421 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3481 accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     3541 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     3601 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     3661 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg
     3721 cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg
     3781 tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg
     3841 ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc
     3901 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
     3961 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
     4021 gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt
     4081 ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct
     4141 gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct
     4201 caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca
     4261 ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg
     4321 atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt
     4381 ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca
     4441 agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag
     4501 agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg
     4561 acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc
     4621 atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca
     4681 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     4741 gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac
     4801 gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac
     4861 gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca
     4921 tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc
     4981 ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc
     5041 agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact
     5101 ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca
     5161 gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg
     5221 ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa
     5281 aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt
     5341 gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc
     5401 actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg
     5461 ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc
     5521 gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga
     5581 ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc
     5641 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
     5701 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
     5761 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
     5821 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
     5881 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
     5941 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
     6001 aagcgct
//
