LOCUS       sequence-343602-        6123 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pET-iBcl2-v4, His tag for affinity purification.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6123)
  AUTHORS   Shui S, Gainza P, Scheller L, Yang C, Kurumida Y, Rosset S, Georgeon
            S, Di Roberto RB, Castellanos-Rueda R, Reddy ST, Correia BE
  TITLE     A rational blueprint for the design of chemically-controlled protein
            switches.
  JOURNAL   Nat Commun. 2021 Oct 1;12(1):5754. doi: 10.1038/s41467-021-25735-9.
  PUBMED    34599176
REFERENCE   2  (bases 1 to 6123)
  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"; journalName: "Nat 
            Commun."; date: "2021-10-1"; volume: "12(1):5754. doi"; pages: " 
            10.1038/s41467-021-25735-9"
COMMENT     Sequence Label: S330
FEATURES             Location/Qualifiers
     source          1..6123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(479..1561)
                     /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     1523..1542
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1562..1639)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1845..1864
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1948..1967
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1948..1966
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1967..1991
                     /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             2006..2028
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2549..2566
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2585..2632
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2807..2812,2813..2829,2830..2835))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2862..2880)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2948..3052
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3053..3121,3122..3913)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3271..3290)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4084..4672
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4573..4592
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4826..4843
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4858..4997
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4923..4942)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5083..5105
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5099..5290)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 atggtcgtca tctacctgcc tggacagcat ggcctgcaac gcgggcatcc cgatgccgcc
       61 ggaagcgaga agaatcataa tggggaaggc catccagcct cgcgtcgcga acgccagcaa
      121 gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg
      181 tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc
      241 aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca
      301 gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc
      361 gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
      421 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
      481 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
      541 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga
      601 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
      661 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
      721 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc
      781 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
      841 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
      901 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
      961 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt
     1021 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga
     1081 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag
     1141 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc
     1201 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc
     1261 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg
     1321 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg
     1381 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg
     1441 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg
     1501 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca
     1561 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt
     1621 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
     1681 aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc
     1741 aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa
     1801 caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata
     1861 taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag
     1921 aggatcgaga tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg
     1981 ataacaattc ccctctagaa ataattttgt ttaactttaa gaaggagata tacatatggc
     2041 acatccgggt cggactggat acgacaatag ggagattgtt atgaagtata tacactataa
     2101 gttgtcacag agagggtatg aatgggacgc cggtgacgat gtggaggaga atagaacaga
     2161 agccccagaa ggcaccgaga gcgaggtagt gcacttgacc ctgagacagg ttggggataa
     2221 tttttcccga cggtaccgac gcgatttcgc tgaaatgagc agtcaacttc acctcacccc
     2281 ttttactgcg cgaggcaggt tcgcaaccgt ggttgaagag ttgttccgag atggcgttaa
     2341 ctggggacgc attgtcgcct tcttcgagtt cggtggcgtt atgtgtgtgg aaagcgtcaa
     2401 ccgagaaatg agcccgcttg ttgataatat cgccctgtgg atgacagagt atctcaacag
     2461 acatttgcac acgtggattc aggataatgg cggatgggat gctttcgtag aattgcatgg
     2521 gcctactagt ggagcaggat ccggatggca ccaccaccac caccactgat gagctgagca
     2581 ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg
     2641 aggaactata tccggatatc ccgcaagagg cccggcagta ccggcataac caagcctatg
     2701 cctacagcat ccagggtgac ggtgccgagg atgacgatga gcgcattgtt agatttcata
     2761 cacggtgcct gactgcgtta gcaatttaac tgtgataaac taccgcatta aagcttatcg
     2821 atgataagct gtcaaacatg agaattcttg aagacgaaag ggcctcgtga tacgcctatt
     2881 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     2941 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     3001 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     3061 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     3121 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     3181 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     3241 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtgttga
     3301 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     3361 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     3421 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     3481 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     3541 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgcagc
     3601 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     3661 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     3721 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     3781 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     3841 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     3901 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     3961 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     4021 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     4081 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     4141 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     4201 cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca
     4261 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     4321 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     4381 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     4441 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     4501 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4561 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4621 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4681 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     4741 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     4801 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct
     4861 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcaatg gtgcactctc
     4921 agtacaatct gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg
     4981 actgggtcat ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     5041 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     5101 agaggttttc accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt
     5161 ggtcgtgaag cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct
     5221 ccagaagcgt taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct
     5281 gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga
     5341 tgaaacgaga gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg
     5401 aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc
     5461 agggtcaatg ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc
     5521 atcctgcgat gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac
     5581 tttacgaaac acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc
     5641 agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc
     5701 aaccccgcca gcctagccgg gtcctcaacg acaggagcac gatcatgcgc acccgtggcc
     5761 aggacccaac gctgcccgag atgcgccgcg tgcggctgct ggagatggcg gacgcgatgg
     5821 atatgttctg ccaagggttg gtttgcgcat tcacagttct ccgcaagaat tgattggctc
     5881 caattcttgg agtggtgaat ccgttagcga ggtgccgccg gcttccattc aggtcgaggt
     5941 ggcccggctc catgcaccgc gacgcaacgc ggggaggcag acaaggtata gggcggcgcc
     6001 tacaatccat gccaacccgt tccatgtgct cgccgaggcg gcataaatcg ccgtgacgat
     6061 cagcggtcca atgatcgaag ttaggctggt aagagccgcg agcgatcctt gaagctgtcc
     6121 ctg
//
