LOCUS       sequence-200224-        5307 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of GRIP PDZ4 domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5307)
  AUTHORS   Tonikian R, Zhang Y, Sazinsky SL, Currell B, Yeh JH, Reva B, Held 
            HA, Appleton BA, Evangelista M, Wu Y, Xin X, Chan AC, Seshagiri S, 
            Lasky LA, Sander C, Boone C, Bader GD, Sidhu SS
  TITLE     A specificity map for the PDZ domain family.
  JOURNAL   PLoS Biol. 2008 Sep 30;6(9):e239. doi: 10.1371/journal.pbio.0060239.
  PUBMED    18828675
REFERENCE   2  (bases 1 to 5307)
  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: "PLoS 
            Biol."; date: "2008-09-30"; volume: "6(9):e239. doi"; pages: " 
            10.1371/journal.pbio.0060239"
COMMENT     Sequence Label: GRIP1_PDZ_4
FEATURES             Location/Qualifiers
     source          1..5307
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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             76..729
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     687..709
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             736..759
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1175..1197)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1342..1360)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1428..1532
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1533..1601,1602..2393)
                     /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(1751..1770)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2564..3152
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3053..3072
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3306..3323
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3396..3473
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3474..4556
                     /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(3493..3512)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4605..4610,4611..4628,4629..4635)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4643..4659
                     /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     4667..4683
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4667..4683
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4679..5206
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4699..4716)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4699..4715)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4708..4730)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      121 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      181 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      241 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      301 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      361 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      421 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      481 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
      541 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
      601 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
      661 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
      721 cctccaaaat cggatctgga agttctgttc caggggcccc tgggatctgg attggctggg
      781 caggttgttc acacagaaac cacagaggtt gtgctgacgg cagatcctgt cacaggattt
      841 gggatccaac tgcagggcag tgtgtttgcc acagaaactc tctcttctcc acctctgatt
      901 tcctatatcg aagctgacag cccagcagag agatgtgggg tgctacagat tggagacaga
      961 gtgatggcca tcaatggaat tccaacagaa gacagcacct tcgaagaagc cagtcagctc
     1021 ctccgagact cttcaatcac gagcaaggtc acactggaaa tcgagtttga tgttgcagag
     1081 tctgtcatcc caagtagtgg atgagtcgac tcgagcggcc gcatcgtgac tgactgacga
     1141 tctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     1201 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     1261 cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt
     1321 ataattcttg aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     1381 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     1441 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     1501 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     1561 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     1621 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     1681 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     1741 ttaaagttct gctatgtggc gcggtattat cccgtgttga cgccgggcaa gagcaactcg
     1801 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     1861 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     1921 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     1981 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     2041 ccataccaaa cgacgagcgt gacaccacga tgcctgcagc aatggcaaca acgttgcgca
     2101 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     2161 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     2221 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     2281 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     2341 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     2401 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     2461 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     2521 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     2581 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     2641 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     2701 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     2761 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     2821 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     2881 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     2941 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     3001 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     3061 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     3121 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     3181 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     3241 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     3301 agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta
     3361 cgcatctgtg cggtatttca caccgcataa attccgacac catcgaatgg cgcaaaacct
     3421 ttcgcggtat ggcatgatag cgcccggaag agagtcaatt cagggtggtg aatgtgaaac
     3481 cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg
     3541 tggtgaacca ggccagccac gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg
     3601 cggagctgaa ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc
     3661 tgattggcgt tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga
     3721 ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg
     3781 gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga
     3841 tcattaacta tccgctggat gaccaggatg ccattgctgt ggaagctgcc tgcactaatg
     3901 ttccggcgtt atttcttgat gtctctgacc agacacccat caacagtatt attttctccc
     3961 atgaagacgg tacgcgactg ggcgtggagc atctggtcgc attgggtcac cagcaaatcg
     4021 cgctgttagc gggcccatta agttctgtct cggcgcgtct gcgtctggct ggctggcata
     4081 aatatctcac tcgcaatcaa attcagccga tagcggaacg ggaaggcgac tggagtgcca
     4141 tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc
     4201 tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag tccgggctgc
     4261 gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagacagc tcatgttata
     4321 tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc
     4381 gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac
     4441 tggtgaaaag aaaaaccacg ctagcgccca atacgcaaac cgcctctccc cgcgcgttgg
     4501 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     4561 aacgcaatta atgtaagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     4621 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     4681 gaccatgatt acggattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     4741 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     4801 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgctt
     4861 tgcctggttt ccggcaccag aagcggtgcc ggaaagctgg ctggagtgcg atcttcctga
     4921 ggccgatact gtcgtcgtcc cctcaaactg gcagatgcac ggttacgatg cgcccatcta
     4981 caccaacgtg acctatccca ttacggtcaa tccgccgttt gttcccacgg agaatccgac
     5041 gggttgttac tcgctcacat ttaatgttga tgaaagctgg ctacaggaag gccagacgcg
     5101 aattattttt gatggcgttg gaattagctt atcgactgca cggtgcacca atgcttctgg
     5161 cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca ctgcataatt
     5221 cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt ttgcgccgac atcataacgg
     5281 ttctggcaaa tattctgaaa tgagctg
//
