LOCUS       sequence-191777-        6951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            ARHGAP32-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6951)
  AUTHORS   Teyra J, Huang H, Jain S, Guan X, Dong A, Liu Y, Tempel W, Min J, 
            Tong Y, Kim PM, Bader GD, Sidhu SS
  TITLE     Comprehensive Analysis of the Human SH3 Domain Family Reveals a Wide
            Variety of Non-canonical Specificities.
  JOURNAL   Structure. 2017 Oct 3;25(10):1598-1610.e3. doi: 
            10.1016/j.str.2017.07.017. Epub 2017 Sep 7.
  PUBMED    28890361
REFERENCE   2  (bases 1 to 6951)
  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: 
            "10.1016/j.str.2017.07.017"; journalName: "Structure"; date: 
            "2017-10-3- 3"; volume: "25"; issue: "10"; pages: "1598-1610.e3"
COMMENT     Sequence Label: pHH0103_ARHGAP32-1/1
FEATURES             Location/Qualifiers
     source          1..6951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        37..114
                     /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             115..1197
                     /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(134..153)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1440..1445,1446..1461,1462..1468)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1476..1492
                     /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     1481..1503
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1577..1594
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1610..2263
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2221..2243
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2270..2293
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2306..2323
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2585..2632
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4225..4680)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4362..4383)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4574..4593
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4722..4744)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4830..4970
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4885..4904
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4985..5002)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5156..5743)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5914..6705,6706..6774))
                     /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     6537..6556
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6775..6879)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     join(6947..6951,1..14)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcgtatcacg aggccctttc gtcttcaaga attcccgaca ccatcgaatg gtgcaaaacc
       61 tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt gaatgtgaaa
      121 ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac cgtttcccgc
      181 gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg
      241 gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg
      301 ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg
      361 attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc
      421 ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg
      481 atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc ctgcactaat
      541 gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat tattttctcc
      601 catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca ccagcaaatc
      661 gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc tggctggcat
      721 aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga ctggagtgcc
      781 atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg
      841 ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg
      901 cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag ctcatgttat
      961 atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac cagcgtggac
     1021 cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt gcccgtctca
     1081 ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     1141 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
     1201 caacgcaatt aatgtaagtt agctcactca ttaggcacaa ttctcatgtt tgacagctta
     1261 tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga agctgtggta
     1321 tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca ctcccgttct
     1381 ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga aatgagctgt
     1441 tgacaattaa tcatcggctc gtataatgtg tggaattgtg agcggataac aatttcacac
     1501 aggaaacagc cagtccgttt aggtgttttc acgagcactt caccaacaag gaccatagat
     1561 tatgaaaatc gaagaacacc atcaccatca ccattccagc ggtaagctta tgtcccctat
     1621 actaggttat tggaaaatta agggccttgt gcaacccact cgacttcttt tggaatatct
     1681 tgaagaaaaa tatgaagagc atttgtatga gcgcgatgaa ggtgataaat ggcgaaacaa
     1741 aaagtttgaa ttgggtttgg agtttcccaa tcttccttat tatattgatg gtgatgttaa
     1801 attaacacag tctatggcca tcatacgtta tatagctgac aagcacaaca tgttgggtgg
     1861 ttgtccaaaa gagcgtgcag agatttcaat gcttgaagga gcggttttgg atattagata
     1921 cggtgtttcg agaattgcat atagtaaaga ctttgaaact ctcaaagttg attttcttag
     1981 caagctacct gaaatgctga aaatgttcga agatcgttta tgtcataaaa catatttaaa
     2041 tggtgatcat gtaacccatc ctgacttcat gttgtatgac gctcttgatg ttgttttata
     2101 catggaccca atgtgcctgg atgcgttccc aaaattagtt tgttttaaaa aacgtattga
     2161 agctatccca caaattgata agtacttgaa atccagcaag tatatagcat ggcctttgca
     2221 gggctggcaa gccacgtttg gtggtggcga ccatcctcca aaatcggatc tggaagttct
     2281 gttccagggg cccctgtcca gcggtctggt tccgcgtggt tccggtaccg cggcccagcc
     2341 ggccccggca gtcggtgcag cgcatgttat taaacgttat acggctcgtg ctccggatga
     2401 actgaccctg gaagttggcg atattgtgtc ggtgattgat atgccgccga aagttctgag
     2461 tacctggtgg cgtggcaaac atggttttca ggtcggcctg ttcccgggtc actgcgtgga
     2521 actgatcaac cagaaagtgc cgcaaagcgt tacgaattct gtcccgaaag cggccgcata
     2581 ataaaccgat acaattaaag gctccttttg gagccttttt tttggagatt ttcaacgtga
     2641 aaaaattatt attcgcaatt cctttagttg ttcctttcta ttctcactcc gctgaaactg
     2701 ttgaaagttg tttagcaaaa ccccatacag aaaattcatt tactaacgtc tggaaagacg
     2761 acaaaacttt agatcgttac gctaactatg agggctgtct gtggaatgct acaggcgttg
     2821 tagtttgtac tggtgacgaa actcagtgtc tagctagagt ggcggtggct ctggttccgg
     2881 tgattttgat tatgaaaaga tggcaaacgc taataagggg gctatgaccg aaaatgccga
     2941 tgaaaacgcg ctacagtctg acgctaaagg caaacttgat tctgtcgcta ctgattacgg
     3001 tgctgctatc gatggtttca ttggtgacgt ttccggcctt gctaatggta atggtgctac
     3061 tggtgatttt gctggctcta attcccaaat ggctcaagtc ggtgacggtg ataattcacc
     3121 tttaatgaat aatttccgtc aatatttacc ttccctccct caatcggttg aatgtcgccc
     3181 ttttgtcttt ggcgctggta aaccatatga attttctatt gattgtgaca aaataaactt
     3241 attccgtggt gtctttgcgt ttcttttata tgttgccacc tttatgtatg tattttctac
     3301 gtttgctaac atactgcgta ataaggagtc ttaatcatgc cagttctttt ggctagcgcc
     3361 gccctatacc ttgtctgcct ccccgcgttg cgtcgcggtg catggagccg ggccacctcg
     3421 acctgaatgg aagccggcgg cacctcgcta acggattcac cactccaaga attggagcca
     3481 atcaattctt gcggagaact gtgaatgcgc aaaccaaccc ttggcagaac atatccatcg
     3541 cgtccgccat ctccagcagc cgcacgcggc gcatctcggg cagcgttggg tcctggccac
     3601 gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg ggttgcctta
     3661 ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc tgctgcaaaa
     3721 cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg taaagtctgg
     3781 aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca ggatgctgct
     3841 ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga ccctgagtga
     3901 tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa cgttccagta
     3961 accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt ttcatcggta
     4021 tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc aaacaggaaa
     4081 aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt ctggagaaac
     4141 tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac gaccacgctg
     4201 atgagcttta ccgcaggatc cgcgaaattg taagcgttaa tattttgtta aaattcgcgt
     4261 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     4321 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     4381 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     4441 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     4501 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     4561 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     4621 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     4681 cgcggatcct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc
     4741 ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc
     4801 gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc
     4861 ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     4921 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg
     4981 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5041 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5101 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5161 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5221 acccgacagg actataaaga taccagggtt tccccctgga agctccctcg tgcgctctcc
     5281 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5341 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5401 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     5461 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5521 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5581 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5641 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5701 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5761 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5821 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5881 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     5941 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     6001 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     6061 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6121 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6181 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgctg caggcatcgt
     6241 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6301 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6361 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6421 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6481 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa
     6541 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6601 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6661 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6721 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6781 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6841 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6901 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata g
//
