LOCUS       sequence-192405-        6956 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            VAV2-2/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6956)
  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 6956)
  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_VAV2-2/2
FEATURES             Location/Qualifiers
     source          1..6956
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     22..40
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        63..140
                     /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             141..1223
                     /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(160..179)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1466..1471,1472..1487,1488..1494)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1502..1518
                     /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     1507..1529
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1603..1620
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1636..2289
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2247..2269
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2296..2319
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2332..2349
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2616..2663
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4256..4711)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4393..4414)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4605..4624
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4753..4775)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4861..5001
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4916..4935
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5016..5033)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5187..5774)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5945..6736,6737..6805))
                     /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     6568..6587
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6806..6910)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct tcaagaattc
       61 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
      121 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      181 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      241 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      301 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      361 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      421 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      481 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      541 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      601 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      661 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      721 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      781 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      841 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      901 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
      961 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
     1021 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1081 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1141 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1201 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1261 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     1321 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1381 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     1441 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     1501 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
     1561 gcacttcacc aacaaggacc atagattatg aaaatcgaag aacaccatca ccatcaccat
     1621 tccagcggta agcttatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
     1681 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
     1741 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
     1801 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
     1861 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
     1921 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
     1981 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
     2041 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
     2101 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
     2161 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
     2221 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
     2281 cctccaaaat cggatctgga agttctgttc caggggcccc tgtccagcgg tctggttccg
     2341 cgtggttccg gtaccgcggc ccagccggcc atggcccagg gcgcgctggg tacagcagtt
     2401 gcccgttaca actttgcagc acgtgatatg cgtgaactgt ctctgcgtga aggtgatgtt
     2461 gttcgtatct actcccgtat tggtggtgat caaggttggt ggaaaggtga aaccaacggt
     2521 cgtattggtt ggtttccgtc cacctacgtt gaagaagaag gtgcggccgc atcctggacg
     2581 acaatgtcat tccgattttc gcggccgcat aataaaccga tacaattaaa ggctcctttt
     2641 ggagcctttt ttttggagat tttcaacgtg aaaaaattat tattcgcaat tcctttagtt
     2701 gttcctttct attctcactc cgctgaaact gttgaaagtt gtttagcaaa accccataca
     2761 gaaaattcat ttactaacgt ctggaaagac gacaaaactt tagatcgtta cgctaactat
     2821 gagggctgtc tgtggaatgc tacaggcgtt gtagtttgta ctggtgacga aactcagtgt
     2881 ctagctagag tggcggtggc tctggttccg gtgattttga ttatgaaaag atggcaaacg
     2941 ctaataaggg ggctatgacc gaaaatgccg atgaaaacgc gctacagtct gacgctaaag
     3001 gcaaacttga ttctgtcgct actgattacg gtgctgctat cgatggtttc attggtgacg
     3061 tttccggcct tgctaatggt aatggtgcta ctggtgattt tgctggctct aattcccaaa
     3121 tggctcaagt cggtgacggt gataattcac ctttaatgaa taatttccgt caatatttac
     3181 cttccctccc tcaatcggtt gaatgtcgcc cttttgtctt tggcgctggt aaaccatatg
     3241 aattttctat tgattgtgac aaaataaact tattccgtgg tgtctttgcg tttcttttat
     3301 atgttgccac ctttatgtat gtattttcta cgtttgctaa catactgcgt aataaggagt
     3361 cttaatcatg ccagttcttt tggctagcgc cgccctatac cttgtctgcc tccccgcgtt
     3421 gcgtcgcggt gcatggagcc gggccacctc gacctgaatg gaagccggcg gcacctcgct
     3481 aacggattca ccactccaag aattggagcc aatcaattct tgcggagaac tgtgaatgcg
     3541 caaaccaacc cttggcagaa catatccatc gcgtccgcca tctccagcag ccgcacgcgg
     3601 cgcatctcgg gcagcgttgg gtcctggcca cgggtgcgca tgatcgtgct cctgtcgttg
     3661 aggacccggc taggctggcg gggttgcctt actggttagc agaatgaatc accgatacgc
     3721 gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg
     3781 gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt
     3841 atgttccgga tctgcatcgc aggatgctgc tggctaccct gtggaacacc tacatctgta
     3901 ttaacgaagc gctggcattg accctgagtg atttttctct ggtcccgccg catccatacc
     3961 gccagttgtt taccctcaca acgttccagt aaccgggcat gttcatcatc agtaacccgt
     4021 atcgtgagca tcctctctcg tttcatcggt atcattaccc ccatgaacag aaatccccct
     4081 tacacggagg catcagtgac caaacaggaa aaaaccgccc ttaacatggc ccgctttatc
     4141 agaagccaga cattaacgct tctggagaaa ctcaacgagc tggacgcgga tgaacaggca
     4201 gacatctgtg aatcgcttca cgaccacgct gatgagcttt accgcaggat ccgcgaaatt
     4261 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     4321 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     4381 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     4441 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     4501 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     4561 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     4621 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     4681 gccgcgctta atgcgccgct acagggcgcg tcgcggatcc tgcctcgcgc gtttcggtga
     4741 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     4801 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     4861 cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca
     4921 tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta
     4981 aggagaaaat accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5041 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5101 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5161 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5221 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccagggt
     5281 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     5341 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5401 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5461 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5521 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5581 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     5641 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5701 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5761 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5821 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5881 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5941 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     6001 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     6061 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     6121 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     6181 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     6241 gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     6301 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccata ttgtgcaaaa
     6361 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6421 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6481 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6541 gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga actttaaaag
     6601 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6661 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6721 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6781 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6841 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6901 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attatt
//
