LOCUS       sequence-149975-        6409 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To purify GST-tagged MKRN1 recombinant protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6409)
  AUTHORS   Lee EW, Lee MS, Camus S, Ghim J, Yang MR, Oh W, Ha NC, Lane DP, Song
            J
  TITLE     Differential regulation of p53 and p21 by MKRN1 E3 ligase controls 
            cell cycle arrest and apoptosis.
  JOURNAL   EMBO J. 2009 Jul 22;28(14):2100-13. doi: 10.1038/emboj.2009.164. 
            Epub 2009 Jun 18.
  PUBMED    19536131
REFERENCE   2  (bases 1 to 6409)
  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: 
            "10.1038/emboj.2009.164"; journalName: "EMBO J"; date: "2009-07-22- 
            22"; volume: "28"; issue: "14"; pages: "2100-13"
COMMENT     Sequence Label: pGEX 4T-1/MKRN1
FEATURES             Location/Qualifiers
     source          1..6409
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2459..2481)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2626..2644)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2712..2816
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2817..2885,2886..3677)
                     /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(3035..3054)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3848..4436
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4337..4356
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4590..4607
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4680..4757
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4758..5840
                     /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     complement(4777..4796)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5853..5874
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5889..5894,5895..5912,5913..5919)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5927..5943
                     /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)."
     primer_bind     5951..5967
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5951..5967
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5963..6409,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5983..6000)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5983..5999)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5992..6014)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatccccgga attcatggcg gaggctgcaa
      961 ctcccggaac aacagccaca acatcaggag caggagcggc agcggcgacg gcggcagcag
     1021 cctcccccac cccgatcccc acagtcaccg ccccgtccct gggggcgggc ggagggggcg
     1081 gcggcagcga cggcagcggc ggcggctgga ctaaacaggt cacctgcagg tattttatgc
     1141 atggggtttg taaggaagga gacaactgtc gctactcgca tgacctctct gacagtccgt
     1201 atagtgtagt gtgcaagtat tttcagcgag ggtactgtat ttatggagac cgctgcagat
     1261 atgaacatag caaaccattg aaacaggaag aagcaactgc tacagagcta actacaaagt
     1321 catcccttgc tgcttcctca agtctctcat cgatagttgg accacttgtt gaaatgaata
     1381 caggcgaagc tgagtcaaga aattcaaact ttgcaactgt aggagcaggt tcagaggact
     1441 gggtgaatgc tattgagttt gttcctgggc aaccctactg tggccgtact gcgccttcct
     1501 gcactgaagc acccctgcag ggctcagtga ccaaggaaga atcagagaaa gagcaaaccg
     1561 ccgtggagac aaagaagcag ctgtgcccct atgctgcagt gggagagtgc cgatacgggg
     1621 agaactgtgt gtatctccac ggagattctt gtgacatgtg tgggctgcag gtcctgcatc
     1681 caatggatgc tgcccagaga tcgcagcata tcaaatcgtg cattgaggcc catgagaagg
     1741 acatggagct ctcatttgcc gtgcagcgca gcaaggacat ggtgtgtggg atctgcatgg
     1801 aggtggtcta tgagaaagcc aaccccagtg agcgccgctt cgggatcctc tccaactgca
     1861 accacaccta ctgtctcaag tgcattcgca agtggaggag tgctaagcaa tttgagagca
     1921 agatcataaa gtcctgccca gaatgccgga tcacatctaa ctttgtcatt ccaagtgagt
     1981 actgggtgga ggagaaagaa gagaagcaga aactcattct gaaatacaag gaggcaatga
     2041 gcaacaaggc gtgcaggtat tttgatgaag gacgtgggag ctgcccattt ggagggaact
     2101 gtttttacaa gcatgcgtac cctgatggcc gtagagagga gccacagaga cagaaagtgg
     2161 gaacatcaag cagataccgg gcccaacgaa ggaaccactt ctgggaactc attgaggaaa
     2221 gagagaacag caaccccttt gacaacgatg aagaagaggt tgtcaccttt gagctgggcg
     2281 agatgttgct tatgcttttg gctgcaggtg gggacgacga actaacagac tctgaagatg
     2341 agtgggactt gtttcatgat gagctggaag atttttatga cttggatcta tagctcgagc
     2401 ggccgcatcg tgactgactg acgatctgcc tcgcgcgttt cggtgatgac ggtgaaaacc
     2461 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     2521 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc
     2581 agtcacgtag cgatagcgga gtgtataatt cttgaagacg aaagggcctc gtgatacgcc
     2641 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     2701 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2761 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     2821 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     2881 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     2941 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     3001 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     3061 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     3121 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     3181 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     3241 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     3301 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     3361 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     3421 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     3481 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     3541 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     3601 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     3661 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     3721 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     3781 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     3841 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     3901 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     3961 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     4021 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     4081 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     4141 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     4201 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     4261 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     4321 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     4381 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     4441 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     4501 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     4561 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     4621 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc ataaattccg
     4681 acaccatcga atggtgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc
     4741 aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg
     4801 tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc
     4861 gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac
     4921 aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg
     4981 cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg
     5041 tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc
     5101 tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg
     5161 ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac
     5221 ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg gagcatctgg
     5281 tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc
     5341 gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg
     5401 aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg
     5461 agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc
     5521 gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg
     5581 ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc
     5641 tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg
     5701 gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg cccaatacgc
     5761 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     5821 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca
     5881 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     5941 caatttcaca caggaaacag ctatgaccat gattacggat tcactggccg tcgttttaca
     6001 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     6061 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     6121 cagcctgaat ggcgaatggc gctttgcctg gtttccggca ccagaagcgg tgccggaaag
     6181 ctggctggag tgcgatcttc ctgaggccga tactgtcgtc gtcccctcaa actggcagat
     6241 gcacggttac gatgcgccca tctacaccaa cgtaacctat cccattacgg tcaatccgcc
     6301 gtttgttccc acggagaatc cgacgggttg ttactcgctc acatttaatg ttgatgaaag
     6361 ctggctacag gaaggccaga cgcgaattat ttttgatggc gttggaatt
//
