LOCUS       sequence-389067-        5855 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of MVPI-SMAC(delta(55))_9His.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5855)
  AUTHORS   Hunkeler M, Jin CY, Fischer ES
  TITLE     Structures of BIRC6-client complexes provide a mechanism of 
            SMAC-mediated release of caspases.
  JOURNAL   Science. 2023 Mar 17;379(6637):1105-1111. doi: 
            10.1126/science.ade5750. Epub 2023 Feb 9.
  PUBMED    36758104
REFERENCE   2  (bases 1 to 5855)
  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: 
            "10.1126/science.ade5750"; journalName: "Science"; date: 
            "2023-03-17- 17"; volume: "379"; issue: "6637"; pages: "1105-1111"
COMMENT     Sequence Label: pNIC-Bio2_MVPI_SMAC(delta(55))_9His
FEATURES             Location/Qualifiers
     source          1..5855
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             641..667
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     CDS             720..737
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(790..808)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      804..851
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      888..1343
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(975..994)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1185..1206
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1435..2250)
                     /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(1435..2250)
                     /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     2158..2177
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2242..2261
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2372..2960
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2861..2880
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3114..3131
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3146..3285
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3211..3230)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3371..3393
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3387..3578)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4387..5469)
                     /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
                     EACKAAVHNLLAQRVSGLIINYQLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5431..5450
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5470..5547)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5753..5772
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat ggttcctatc gcgcagaaat ctgaacctca
      121 ttccttgtcg tcggaagctt tgatgcgtcg tgctgtatct ttagtcacgg attccacctc
      181 aacttttttg agccaaacaa catatgcgct tattgaagcg attacagaat acactaaagc
      241 ggtgtatact ctgactagtt tgtatcgcca gtatacttcc cttcttggaa aaatgaatag
      301 tgaagaagag gatgaagtat ggcaagttat tatcggagcc cgcgcggaaa tgactagtaa
      361 acatcaggag tacttgaagt tagaaaccac ctggatgacc gctgtagggt tgagtgagat
      421 ggctgctgaa gcggcctatc agactggagc cgatcaggca tcaattacag cacgtaacca
      481 catccaatta gtgaagctgc aagtcgaaga agtacaccaa ctttcgcgca aagcagagac
      541 caaattggcc gaggctcaaa ttgaggagct gcgccaaaaa acccaggaag agggggaaga
      601 acgtgcagaa tcggagcagg aggcttatct tcgtgaggat caccatcatc atcatcatca
      661 ccatcattga taaggatccg aattcgagct ccgtcgacaa gcttgcggcc gcactcgagc
      721 accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
      781 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
      841 ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag
      901 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
      961 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     1021 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     1081 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     1141 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     1201 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     1261 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     1321 caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct
     1381 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgaatta attcttagaa
     1441 aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata
     1501 tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat
     1561 ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa
     1621 tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc
     1681 cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag gccagccatt
     1741 acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg
     1801 agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa
     1861 ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc
     1921 taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg
     1981 agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct
     2041 gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc
     2101 tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc
     2161 gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctaga
     2221 gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc
     2281 agacagtttt attgttcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     2341 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     2401 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     2461 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     2521 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     2581 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     2641 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     2701 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     2761 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     2821 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     2881 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     2941 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     3001 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     3061 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     3121 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     3181 gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     3241 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
     3301 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     3361 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     3421 aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt
     3481 tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag
     3541 cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga
     3601 tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt
     3661 actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg
     3721 atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat
     3781 gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg
     3841 cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat
     3901 gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc
     3961 ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac
     4021 aggagcacga tcatgcgcac ccgtggggcc gccatgccgg cgataatggc ctgcttctcg
     4081 ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg caagattccg
     4141 aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa
     4201 atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa gacagtcata
     4261 agtgcggcga cgatagtcat gtcccgcgcc caccggaagg agctgactgg gttgaaggct
     4321 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     4381 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4441 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     4501 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     4561 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     4621 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     4681 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     4741 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     4801 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     4861 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     4921 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     4981 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     5041 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagct gatagttaat
     5101 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     5161 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     5221 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     5281 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     5341 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     5401 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     5461 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     5521 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     5581 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     5641 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
     5701 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
     5761 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
     5821 cggcgtagag gatcgagatc tcgatcccgc gaaat
//
