LOCUS       sequence-192279-        6912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            ARHGEF9-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6912)
  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 6912)
  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_ARHGEF9-1/1
FEATURES             Location/Qualifiers
     source          1..6912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     8..26
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        49..126
                     /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             127..1209
                     /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(146..165)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1452..1457,1458..1473,1474..1480)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1488..1504
                     /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     1493..1515
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1589..1606
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1622..2275
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2233..2255
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2282..2305
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2318..2335
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2558..2605
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4198..4653)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4335..4356)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4547..4566
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4695..4717)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4803..4943
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4858..4877
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4958..4975)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5129..5716)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5887..6678,6679..6747))
                     /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     6510..6529
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6748..6852)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctataaaaat aggcgtatca cgaggccctt tcgtcttcaa gaattcccga caccatcgaa
       61 tggtgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
      121 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
      181 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
      241 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
      301 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
      361 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
      421 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
      481 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
      541 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
      601 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
      661 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
      721 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
      781 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
      841 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
      901 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac
      961 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
     1021 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
     1081 ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
     1141 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     1201 gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac aattctcatg
     1261 tttgacagct tatcatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg
     1321 gaagctgtgg tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg
     1381 cactcccgtt ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct
     1441 gaaatgagct gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata
     1501 acaatttcac acaggaaaca gccagtccgt ttaggtgttt tcacgagcac ttcaccaaca
     1561 aggaccatag attatgaaaa tcgaagaaca ccatcaccat caccattcca gcggtaagct
     1621 tatgtcccct atactaggtt attggaaaat taagggcctt gtgcaaccca ctcgacttct
     1681 tttggaatat cttgaagaaa aatatgaaga gcatttgtat gagcgcgatg aaggtgataa
     1741 atggcgaaac aaaaagtttg aattgggttt ggagtttccc aatcttcctt attatattga
     1801 tggtgatgtt aaattaacac agtctatggc catcatacgt tatatagctg acaagcacaa
     1861 catgttgggt ggttgtccaa aagagcgtgc agagatttca atgcttgaag gagcggtttt
     1921 ggatattaga tacggtgttt cgagaattgc atatagtaaa gactttgaaa ctctcaaagt
     1981 tgattttctt agcaagctac ctgaaatgct gaaaatgttc gaagatcgtt tatgtcataa
     2041 aacatattta aatggtgatc atgtaaccca tcctgacttc atgttgtatg acgctcttga
     2101 tgttgtttta tacatggacc caatgtgcct ggatgcgttc ccaaaattag tttgttttaa
     2161 aaaacgtatt gaagctatcc cacaaattga taagtacttg aaatccagca agtatatagc
     2221 atggcctttg cagggctggc aagccacgtt tggtggtggc gaccatcctc caaaatcgga
     2281 tctggaagtt ctgttccagg ggcccctgtc cagcggtctg gttccgcgtg gttccggtac
     2341 cgcggcccag ccggccatgg cccagggcgc gctggtttct gcagaagccg tttgggatca
     2401 tgttaccatg gccaatcgtg aactggcatt caaagccggt gatgtcatca aagttctgga
     2461 tgcctccaac aaagattggt ggtggggtca gattgatgat gaagaaggtt ggtttccggc
     2521 atcttttgtt cgtctgtggg ttgcggccgc ataataaacc gatacaatta aaggctcctt
     2581 ttggagcctt ttttttggag attttcaacg tgaaaaaatt attattcgca attcctttag
     2641 ttgttccttt ctattctcac tccgctgaaa ctgttgaaag ttgtttagca aaaccccata
     2701 cagaaaattc atttactaac gtctggaaag acgacaaaac tttagatcgt tacgctaact
     2761 atgagggctg tctgtggaat gctacaggcg ttgtagtttg tactggtgac gaaactcagt
     2821 gtctagctag agtggcggtg gctctggttc cggtgatttt gattatgaaa agatggcaaa
     2881 cgctaataag ggggctatga ccgaaaatgc cgatgaaaac gcgctacagt ctgacgctaa
     2941 aggcaaactt gattctgtcg ctactgatta cggtgctgct atcgatggtt tcattggtga
     3001 cgtttccggc cttgctaatg gtaatggtgc tactggtgat tttgctggct ctaattccca
     3061 aatggctcaa gtcggtgacg gtgataattc acctttaatg aataatttcc gtcaatattt
     3121 accttccctc cctcaatcgg ttgaatgtcg cccttttgtc tttggcgctg gtaaaccata
     3181 tgaattttct attgattgtg acaaaataaa cttattccgt ggtgtctttg cgtttctttt
     3241 atatgttgcc acctttatgt atgtattttc tacgtttgct aacatactgc gtaataagga
     3301 gtcttaatca tgccagttct tttggctagc gccgccctat accttgtctg cctccccgcg
     3361 ttgcgtcgcg gtgcatggag ccgggccacc tcgacctgaa tggaagccgg cggcacctcg
     3421 ctaacggatt caccactcca agaattggag ccaatcaatt cttgcggaga actgtgaatg
     3481 cgcaaaccaa cccttggcag aacatatcca tcgcgtccgc catctccagc agccgcacgc
     3541 ggcgcatctc gggcagcgtt gggtcctggc cacgggtgcg catgatcgtg ctcctgtcgt
     3601 tgaggacccg gctaggctgg cggggttgcc ttactggtta gcagaatgaa tcaccgatac
     3661 gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc gacctgagca acaacatgaa
     3721 tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca
     3781 ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca cctacatctg
     3841 tattaacgaa gcgctggcat tgaccctgag tgatttttct ctggtcccgc cgcatccata
     3901 ccgccagttg tttaccctca caacgttcca gtaaccgggc atgttcatca tcagtaaccc
     3961 gtatcgtgag catcctctct cgtttcatcg gtatcattac ccccatgaac agaaatcccc
     4021 cttacacgga ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg gcccgcttta
     4081 tcagaagcca gacattaacg cttctggaga aactcaacga gctggacgcg gatgaacagg
     4141 cagacatctg tgaatcgctt cacgaccacg ctgatgagct ttaccgcagg atccgcgaaa
     4201 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     4261 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     4321 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     4381 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     4441 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     4501 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     4561 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     4621 ccgccgcgct taatgcgccg ctacagggcg cgtcgcggat cctgcctcgc gcgtttcggt
     4681 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     4741 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     4801 ggcgcagcca tgacccagtc acgtagcgat agcggagtgt atactggctt aactatgcgg
     4861 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     4921 taaggagaaa ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4981 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     5041 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5101 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     5161 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5221 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5281 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5341 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5401 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5461 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5521 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     5581 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5641 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5701 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5761 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5821 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5881 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5941 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6001 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6061 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6121 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6181 gcgcaacgtt gttgccattg ctgcaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6241 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tattgtgcaa
     6301 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6361 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6421 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6481 gagttgctct tgcccggcgt caacacggga taataccgcg ccacatagca gaactttaaa
     6541 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6601 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6661 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6721 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6781 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6841 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     6901 catgacatta ac
//
