LOCUS       sequence-427380-        5594 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5594)
  AUTHORS   Bagrodia S, Taylor SJ, Creasy CL, Chernoff J, Cerione RA
  TITLE     Identification of a mouse p21Cdc42/Rac activated kinase.
  JOURNAL   J Biol Chem. 1995 Sep 29. 270(39):22731-7.
  PUBMED    7559398
REFERENCE   2  (bases 1 to 5594)
  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"; journalName: "J Biol 
            Chem. 1995 Sep 29. 270(39):22731-7."
COMMENT     Sequence Label: pGEX-RhoA
FEATURES             Location/Qualifiers
     source          1..5594
                     /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"
     CDS             936..1517
                     /codon_start=1
                     /product="human Rho family small GTPase RhoA"
                     /label=RhoA
                     /translation="MAAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVAD
                     IEVDGKQVELALWDTAGQEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVK
                     HFCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKT
                     KDGVREVFEMATRAALQARRGKKKSGCLVL"
     primer_bind     complement(1644..1666)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1811..1829)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1897..2001
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2002..2070,2071..2862)
                     /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(2220..2239)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3033..3621
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3522..3541
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3775..3792
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3865..3942
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3943..5025
                     /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(3962..3981)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5074..5079,5080..5097,5098..5104)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5112..5128
                     /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     5136..5152
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5136..5152
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5148..5594,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5168..5185)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5168..5184)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5177..5199)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga 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 gatccatggc tgccatccgg aagaaactgg
      961 tgattgttgg tgatggagcc tgtggaaaga catgcttgct catagtcttc agcaaggacc
     1021 agttcccaga ggtgtatgtg cccacagtgt ttgagaacta tgtggcagat atcgaggtgg
     1081 atggaaagca ggtagagttg gctttgtggg acacagctgg gcaggaagat tatgatcgcc
     1141 tgaggcccct ctcctaccca gataccgatg ttatactgat gtgtttttcc atcgacagcc
     1201 ctgatagttt agaaaacatc ccagaaaagt ggaccccaga agtcaagcat ttctgtccca
     1261 acgtgcccat catcctggtt gggaataaga aggatcttcg gaatgatgag cacacaaggc
     1321 gggagctagc caagatgaag caggagccgg tgaaacctga agaaggcaga gatatggcaa
     1381 acaggattgg cgcttttggg tacatggagt gttcagcaaa gaccaaagat ggagtgagag
     1441 aggtttttga aatggctacg agagctgctc tgcaagctag acgtgggaag aaaaaatctg
     1501 ggtgccttgt cttgtgaaac cttgctggaa gcacagccct tatgcggtga attctagact
     1561 ccatgggtcg actcgagctc aagcttaatt catcgtgact gactgacgat ctgcctcgcg
     1621 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     1681 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     1741 gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta taattcttga
     1801 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
     1861 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     1921 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     1981 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     2041 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     2101 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     2161 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     2221 ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata
     2281 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     2341 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     2401 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     2461 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     2521 gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact
     2581 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     2641 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     2701 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     2761 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     2821 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     2881 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     2941 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     3001 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     3061 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3121 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     3181 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3241 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3301 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3361 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3421 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3481 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     3541 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3601 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3661 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     3721 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     3781 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     3841 ggtatttcac accgcataaa ttccgacacc atcgaatggc gcaaaacctt tcgcggtatg
     3901 gcatgatagc gcccggaaga gagtcaattc agggtggtga atgtgaaacc agtaacgtta
     3961 tacgatgtcg cagagtatgc cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag
     4021 gccagccacg tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc ggagctgaat
     4081 tacattccca accgcgtggc acaacaactg gcgggcaaac agtcgttgct gattggcgtt
     4141 gccacctcca gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc
     4201 gccgatcaac tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc
     4261 tgtaaagcgg cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat cattaactat
     4321 ccgctggatg accaggatgc cattgctgtg gaagctgcct gcactaatgt tccggcgtta
     4381 tttcttgatg tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt
     4441 acgcgactgg gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg
     4501 ggcccattaa gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact
     4561 cgcaatcaaa ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt
     4621 caacaaacca tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac
     4681 gatcagatgg cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg
     4741 gatatctcgg tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtta
     4801 accaccatca aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa
     4861 ctctctcagg gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga
     4921 aaaaccaccc tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     4981 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     5041 tgtaagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     5101 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta
     5161 cggattcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     5221 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     5281 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcttt gcctggtttc
     5341 cggcaccaga agcggtgccg gaaagctggc tggagtgcga tcttcctgag gccgatactg
     5401 tcgtcgtccc ctcaaactgg cagatgcacg gttacgatgc gcccatctac accaacgtga
     5461 cctatcccat tacggtcaat ccgccgtttg ttcccacgga gaatccgacg ggttgttact
     5521 cgctcacatt taatgttgat gaaagctggc tacaggaagg ccagacgcga attatttttg
     5581 atggcgttgg aatt
//
