LOCUS       sequence-191069-        6897 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            VAV1-2/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6897)
  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 6897)
  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_VAV1-2/2
FEATURES             Location/Qualifiers
     source          1..6897
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..20
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        43..120
                     /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             121..1203
                     /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(140..159)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1446..1451,1452..1467,1468..1474)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1482..1498
                     /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     1487..1509
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1583..1600
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1616..2269
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2227..2249
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2276..2299
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2312..2329
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2537..2584
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4177..4632)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4314..4335)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4526..4545
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4674..4696)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4782..4922
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4837..4856
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4937..4954)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5108..5695)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5866..6657,6658..6726))
                     /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     6489..6508
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6727..6831)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaataggcgt atcacgaggc cctttcgtct tcaagaattc ccgacaccat cgaatggtgc
       61 aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat
      121 gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt
      181 tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg
      241 gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag
      301 tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc
      361 gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa
      421 cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt
      481 gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc
      541 actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt
      601 ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag
      661 caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc
      721 tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg
      781 agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact
      841 gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc
      901 gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga agacagctca
      961 tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc
     1021 gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc
     1081 gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc
     1141 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag
     1201 tgagcgcaac gcaattaatg taagttagct cactcattag gcacaattct catgtttgac
     1261 agcttatcat cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc atcggaagct
     1321 gtggtatggc tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc
     1381 cgttctggat aatgtttttt gcgccgacat cataacggtt ctggcaaata ttctgaaatg
     1441 agctgttgac aattaatcat cggctcgtat aatgtgtgga attgtgagcg gataacaatt
     1501 tcacacagga aacagccagt ccgtttaggt gttttcacga gcacttcacc aacaaggacc
     1561 atagattatg aaaatcgaag aacaccatca ccatcaccat tccagcggta agcttatgtc
     1621 ccctatacta ggttattgga aaattaaggg ccttgtgcaa cccactcgac ttcttttgga
     1681 atatcttgaa gaaaaatatg aagagcattt gtatgagcgc gatgaaggtg ataaatggcg
     1741 aaacaaaaag tttgaattgg gtttggagtt tcccaatctt ccttattata ttgatggtga
     1801 tgttaaatta acacagtcta tggccatcat acgttatata gctgacaagc acaacatgtt
     1861 gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt gaaggagcgg ttttggatat
     1921 tagatacggt gtttcgagaa ttgcatatag taaagacttt gaaactctca aagttgattt
     1981 tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat cgtttatgtc ataaaacata
     2041 tttaaatggt gatcatgtaa cccatcctga cttcatgttg tatgacgctc ttgatgttgt
     2101 tttatacatg gacccaatgt gcctggatgc gttcccaaaa ttagtttgtt ttaaaaaacg
     2161 tattgaagct atcccacaaa ttgataagta cttgaaatcc agcaagtata tagcatggcc
     2221 tttgcagggc tggcaagcca cgtttggtgg tggcgaccat cctccaaaat cggatctgga
     2281 agttctgttc caggggcccc tgtccagcgg tctggttccg cgtggttccg gtaccgcggc
     2341 ccagccggcc ggcaccgcca aagcccgtta tgacttctgc gcccgtgacc gctccgaact
     2401 gtccctgaaa gaaggtgaca ttatcaaaat cctgaataaa aaaggccagc aaggttggtg
     2461 gcgtggcgaa atttatggcc gcgtgggttg gtttccggcg aactatgttg aagaagatta
     2521 cgcggccgca taataaaccg atacaattaa aggctccttt tggagccttt tttttggaga
     2581 ttttcaacgt gaaaaaatta ttattcgcaa ttcctttagt tgttcctttc tattctcact
     2641 ccgctgaaac tgttgaaagt tgtttagcaa aaccccatac agaaaattca tttactaacg
     2701 tctggaaaga cgacaaaact ttagatcgtt acgctaacta tgagggctgt ctgtggaatg
     2761 ctacaggcgt tgtagtttgt actggtgacg aaactcagtg tctagctaga gtggcggtgg
     2821 ctctggttcc ggtgattttg attatgaaaa gatggcaaac gctaataagg gggctatgac
     2881 cgaaaatgcc gatgaaaacg cgctacagtc tgacgctaaa ggcaaacttg attctgtcgc
     2941 tactgattac ggtgctgcta tcgatggttt cattggtgac gtttccggcc ttgctaatgg
     3001 taatggtgct actggtgatt ttgctggctc taattcccaa atggctcaag tcggtgacgg
     3061 tgataattca cctttaatga ataatttccg tcaatattta ccttccctcc ctcaatcggt
     3121 tgaatgtcgc ccttttgtct ttggcgctgg taaaccatat gaattttcta ttgattgtga
     3181 caaaataaac ttattccgtg gtgtctttgc gtttctttta tatgttgcca cctttatgta
     3241 tgtattttct acgtttgcta acatactgcg taataaggag tcttaatcat gccagttctt
     3301 ttggctagcg ccgccctata ccttgtctgc ctccccgcgt tgcgtcgcgg tgcatggagc
     3361 cgggccacct cgacctgaat ggaagccggc ggcacctcgc taacggattc accactccaa
     3421 gaattggagc caatcaattc ttgcggagaa ctgtgaatgc gcaaaccaac ccttggcaga
     3481 acatatccat cgcgtccgcc atctccagca gccgcacgcg gcgcatctcg ggcagcgttg
     3541 ggtcctggcc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc
     3601 ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact
     3661 gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt
     3721 cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg
     3781 caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt
     3841 gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac
     3901 aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc
     3961 gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga
     4021 ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc
     4081 ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc
     4141 acgaccacgc tgatgagctt taccgcagga tccgcgaaat tgtaagcgtt aatattttgt
     4201 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     4261 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     4321 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     4381 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     4441 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     4501 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     4561 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     4621 tacagggcgc gtcgcggatc ctgcctcgcg cgtttcggtg atgacggtga aaacctctga
     4681 cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa
     4741 gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca
     4801 cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag attgtactga
     4861 gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     4921 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     4981 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5041 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5101 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5161 agaggtggcg aaacccgaca ggactataaa gataccaggg tttccccctg gaagctccct
     5221 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5281 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5341 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5401 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5461 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5521 gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc
     5581 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5641 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     5701 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     5761 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     5821 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     5881 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     5941 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6001 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6061 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6121 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6181 tgcaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6241 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6301 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6361 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6421 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6481 aacacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6541 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6601 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     6661 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     6721 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     6781 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     6841 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataa
//
