LOCUS       sequence-433888-        6646 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of  GST-tagged CIP2A_1-560 with L533E mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6646)
  TITLE     Westermarck Lab CIP2A Plasmids 2024
REFERENCE   2  (bases 1 to 6646)
  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"
COMMENT     Sequence Label: pGEX-4T_CIP2A_1-560_L533E
FEATURES             Location/Qualifiers
     source          1..6646
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(182..187,188..203,204..210)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    218..234
                     /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             257..910
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     868..890
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             917..934
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2695..2717)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     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"
     promoter        4916..4993
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4994..6076
                     /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(5013..5032)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6125..6130,6131..6148,6149..6155)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6163..6179
                     /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     6187..6203
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6187..6203
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(6199..6646,1..80)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6219..6236)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6219..6235)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6228..6250)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gcttatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg
       61 tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt
      121 ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct
      181 gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata acaatttcac
      241 acaggaaaca gtattcatgt cccctatact aggttattgg aaaattaagg gccttgtgca
      301 acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg
      361 cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct
      421 tccttattat attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat
      481 agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct
      541 tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt
      601 tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga
      661 tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt
      721 gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa
      781 attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc
      841 cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca
      901 tcctccaaaa tcggatctgg ttccgcgtgg atccccagga attcccatgg actccactgc
      961 ctgcttgaag tccttgctcc tgactgtcag tcagtacaaa gccgtgaagt cagaggcgaa
     1021 cgccactcag cttttgcggc acttggaggt aatttctgga cagaaactca cacgactatt
     1081 tacatcaaat cagatattaa caagtgaatg cttgagttgc cttgtagagc tacttgaaga
     1141 ccccaacata agtgcttcac tgatcttaag tattatcggt ttgctgtctc aactagcagt
     1201 agacattgaa accagagatt gtcttcagaa tacatataat ctgaatagtg tgctggcggg
     1261 agtggtttgt cggagcagcc acactgattc ggtgtttttg cagtgcattc aacttctaca
     1321 gaagttaaca tataatgtca aaattttcta ttctggtgcc aatatagatg aattaattac
     1381 gttcctgata gatcacattc aatcttctga agatgagtta aaaatgcctt gtctaggatt
     1441 attggcaaat ctttgtcggc acaatctttc tgttcaaacg cacataaaga cattgagtaa
     1501 tgtgaaatct ttttatcgaa ctcttatcac cttgttggcc catagtagtt taactgtggt
     1561 tgtgtttgca ctttcaatat tatccagttt gacattaaat gaagaggtgg gggaaaagct
     1621 attccatgct cgaaacattc atcagacttt tcaactaata tttaatattc tcataaacgg
     1681 tgatggcact ctaactagaa agtattcagt tgacctactg atggatctcc ttaagaatcc
     1741 taaaattgct gattatctca ccagatatga gcacttttct tcatgtcttc accaagtatt
     1801 aggtcttctt aatggaaagg atcctgattc ctcttcaaag gttttagaat tacttcttgc
     1861 cttctgttca gtgactcagc tgcgccatat gctcactcag atgatgtttg aacagtctcc
     1921 acctggcagc gccactctgg gaagccatac taaatgttta gaacctactg tggctctact
     1981 gcgctggtta agccaacctt tggacggatc agaaaactgt tctgttttag cattggagtt
     2041 gttcaaggaa atatttgagg atgtcataga tgctgctaac tgttcctcgg ctgatcgttt
     2101 tgtgaccctt ctgctgccta caatccttga tcaacttcag ttcacagaac aaaatctaga
     2161 tgaggcttta acaagaaaaa aatgtgaaag gattgccaag gccattgaag ttttgttaac
     2221 tctctgtgga gatgatacac taaaaatgca tattgcaaaa atcttgacaa ctgtcaagtg
     2281 taccactctt atagaacaac aatttacata tggcaagatt gacctgggat ttggaacaaa
     2341 ggttgcagat tctgaattat gcaaacttgc tgctgatgta attttgaaaa ctcttgattt
     2401 gattaacaaa cttaaaccat tggttcctgg tatggaagta agcttctaca aaatacttca
     2461 ggacccacgt ttgattactc ctttggcttt tgctttaacg tcagataata gagaacaagt
     2521 acagtctgga ctgagaatat tagaggaggc tgctccactg ccagattttc ctgctttagt
     2581 acttggagaa agtatagcag caaacaatgc ctatagacaa caggaataac tcgagcggcc
     2641 gcatcgtgac tgactgacga tctgcctcgc gcgtttcggt gatgacggtg aaaacctctg
     2701 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     2761 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc
     2821 acgtagcgat agcggagtgt ataattcttg 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 caccgcataa attccgacac
     4921 catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt
     4981 cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc
     5041 ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga
     5101 aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact
     5161 ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc
     5221 gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt
     5281 gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc
     5341 gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt
     5401 ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat
     5461 caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc atctggtcgc
     5521 attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct
     5581 gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg
     5641 ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg
     5701 catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc
     5761 cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac
     5821 cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct
     5881 ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa
     5941 tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac
     6001 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
     6061 ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat taggcacccc
     6121 aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat
     6181 ttcacacagg aaacagctat gaccatgatt acggattcac tggccgtcgt tttacaacgt
     6241 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     6301 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     6361 ctgaatggcg aatggcgctt tgcctggttt ccggcaccag aagcggtgcc ggaaagctgg
     6421 ctggagtgcg atcttcctga ggccgatact gtcgtcgtcc cctcaaactg gcagatgcac
     6481 ggttacgatg cgcccatcta caccaacgtg acctatccca ttacggtcaa tccgccgttt
     6541 gttcccacgg agaatccgac gggttgttac tcgctcacat ttaatgttga tgaaagctgg
     6601 ctacaggaag gccagacgcg aattattttt gatggcgttg gaatta
//
