LOCUS       sequence-147383-        6030 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST tagged GINA3 protein in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6030)
  AUTHORS   Lin C, Ear J, Midde K, Lopez-Sanchez I, Aznar N, Garcia-Marcos M, 
            Kufareva I, Abagyan R, Ghosh P
  TITLE     Structural basis for activation of trimeric Gi proteins by multiple 
            growth factor receptors via GIV/Girdin.
  JOURNAL   Mol Biol Cell. 2014 Nov 5;25(22):3654-71. doi: 
            10.1091/mbc.E14-05-0978. Epub 2014 Sep 3.
  PUBMED    25187647
REFERENCE   2  (bases 1 to 6030)
  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: 
            "10.1091/mbc.E14-05-0978"; journalName: "Mol Biol Cell"; date: 
            "2014-11-5- 5"; volume: "25"; issue: "22"; pages: "3654-71"
COMMENT     Sequence Label: GST-GINA3
FEATURES             Location/Qualifiers
     source          1..6030
                     /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(2080..2102)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2247..2265)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2333..2437
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2438..2506,2507..3298)
                     /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(2656..2675)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3469..4057
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3958..3977
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4211..4228
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4301..4378
                     /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             4379..5461
                     /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(4398..4417)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5474..5495
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5510..5515,5516..5533,5534..5540)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5548..5564
                     /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     5572..5588
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5572..5588
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5584..6030,1..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(5604..5621)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5604..5620)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5613..5635)
                     /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 attcatgggc tgcacgttga
      961 gcgccgggga caaggcggcg gtggagcgga gtaagatgat cgaccgcaac ttgcgggagg
     1021 acggagagaa agcggccaaa gaagtgaagc tgctgctgct cggcgctgga gaatctggta
     1081 aaagtactat tgtgaaacag atgaaaatca ttcatgagga tggctattcc gaggacgagt
     1141 gtaagcagta taaagttgtc gtctacagca ataccattca gtccatcatt gcaatcataa
     1201 gagccatggg acggttgaag attgattttg gggaagctgc cagagcggat gatgcccgac
     1261 agttatttgt tttagctggc agtgctgagg aaggagtcat gacttcagaa ctagcaggcg
     1321 tgattaaacg tttatggcga gatggcgggg tgcaggcatg cttcagcagg tccagggaat
     1381 atcagctcaa tgattctgct tcatattacc taaatgattt ggatagaata tcccagacca
     1441 actacattcc aactcagcaa gatgttcttc ggacgagagt gaagactaca ggcattgtgg
     1501 agacccactt caccttcaag gagctctact tcaaaatgtt tgatgtaggt ggccaaagat
     1561 ccgaacggaa aaagtggatc cactgttttg agggagtgac agcaattatc ttttgtgtgg
     1621 ctctcagtga ttacgacctt gttctggctg aggacgagga aatgaaccga atgcatgaaa
     1681 gcatgaaatt gtttgacagc atttgtaaca acaaatggtt tacagacact tcaatcattc
     1741 tcttccttaa taagaaagac ctttttgagg aaaaaataaa gaggagtcca ttaacaatct
     1801 gttatccaga atacacaggt tccaatacgt acgaagaggc agctgcttac atccagtgcc
     1861 agtttgaaga tctgaaccga agaaaggaca ccaaggaggt ctacactcac tttacctgtg
     1921 ccactgacac caaaaacgtg cagtttgttt ttgatgctgt tacagatgtc atcattaaaa
     1981 acaacttaaa ggaatgtgga ctttattgag tcgactcgag cggccgcatc gtgactgact
     2041 gacgatctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     2101 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     2161 gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg
     2221 agtgtataat tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     2281 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     2341 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2401 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     2461 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     2521 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     2581 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     2641 cacttttaaa gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca
     2701 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     2761 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     2821 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     2881 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     2941 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt
     3001 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     3061 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     3121 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     3181 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     3241 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     3301 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3361 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     3421 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     3481 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     3541 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     3601 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     3661 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     3721 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     3781 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     3841 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     3901 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     3961 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4021 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4081 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4141 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     4201 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct
     4261 ccttacgcat ctgtgcggta tttcacaccg cataaattcc gacaccatcg aatggtgcaa
     4321 aacctttcgc ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt
     4381 gaaaccagta acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc
     4441 ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc
     4501 gatggcggag ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc
     4561 gttgctgatt ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc
     4621 ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg
     4681 aagcggcgtc gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg
     4741 gctgatcatt aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac
     4801 taatgttccg gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt
     4861 ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca
     4921 aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg
     4981 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
     5041 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
     5101 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
     5161 gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg
     5221 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
     5281 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
     5341 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
     5401 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     5461 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     5521 tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca
     5581 gctatgacca tgattacgga ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     5641 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     5701 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     5761 cgctttgcct ggtttccggc accagaagcg gtgccggaaa gctggctgga gtgcgatctt
     5821 cctgaggccg atactgtcgt cgtcccctca aactggcaga tgcacggtta cgatgcgccc
     5881 atctacacca acgtaaccta tcccattacg gtcaatccgc cgtttgttcc cacggagaat
     5941 ccgacgggtt gttactcgct cacatttaat gttgatgaaa gctggctaca ggaaggccag
     6001 acgcgaatta tttttgatgg cgttggaatt
//
