LOCUS       sequence-192241-        6912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            DNMBP-4/6.
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_DNMBP-4/6
FEATURES             Location/Qualifiers
     source          1..6912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     6..24
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        47..124
                     /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             125..1207
                     /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(144..163)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1450..1455,1456..1471,1472..1478)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1486..1502
                     /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     1491..1513
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1587..1604
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1620..2273
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2231..2253
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2280..2303
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2316..2333
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2556..2603
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4196..4651)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4333..4354)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4545..4564
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4693..4715)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4801..4941
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4856..4875
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4956..4973)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5127..5714)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5885..6676,6677..6745))
                     /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     6508..6527
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6746..6850)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga attcccgaca ccatcgaatg
       61 gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt
      121 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac
      181 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga
      241 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa
      301 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat
      361 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt
      421 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt
      481 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc
      541 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat
      601 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca
      661 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc
      721 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga
      781 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc
      841 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga
      901 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag
      961 ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac
     1021 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt
     1081 gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc
     1141 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     1201 gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcacaa ttctcatgtt
     1261 tgacagctta tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga
     1321 agctgtggta tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca
     1381 ctcccgttct ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga
     1441 aatgagctgt tgacaattaa tcatcggctc gtataatgtg tggaattgtg agcggataac
     1501 aatttcacac aggaaacagc cagtccgttt aggtgttttc acgagcactt caccaacaag
     1561 gaccatagat tatgaaaatc gaagaacacc atcaccatca ccattccagc ggtaagctta
     1621 tgtcccctat actaggttat tggaaaatta agggccttgt gcaacccact cgacttcttt
     1681 tggaatatct tgaagaaaaa tatgaagagc atttgtatga gcgcgatgaa ggtgataaat
     1741 ggcgaaacaa aaagtttgaa ttgggtttgg agtttcccaa tcttccttat tatattgatg
     1801 gtgatgttaa attaacacag tctatggcca tcatacgtta tatagctgac aagcacaaca
     1861 tgttgggtgg ttgtccaaaa gagcgtgcag agatttcaat gcttgaagga gcggttttgg
     1921 atattagata cggtgtttcg agaattgcat atagtaaaga ctttgaaact ctcaaagttg
     1981 attttcttag caagctacct gaaatgctga aaatgttcga agatcgttta tgtcataaaa
     2041 catatttaaa tggtgatcat gtaacccatc ctgacttcat gttgtatgac gctcttgatg
     2101 ttgttttata catggaccca atgtgcctgg atgcgttccc aaaattagtt tgttttaaaa
     2161 aacgtattga agctatccca caaattgata agtacttgaa atccagcaag tatatagcat
     2221 ggcctttgca gggctggcaa gccacgtttg gtggtggcga ccatcctcca aaatcggatc
     2281 tggaagttct gttccagggg cccctgtcca gcggtctggt tccgcgtggt tccggtaccg
     2341 cggcccagcc ggccatggcc cagggcgcgc tgacctatgg tgttgccctg tatcgttttc
     2401 aagccctgga accgaacgaa ctggatttcg aagttggtga caaaatccgt attctggcca
     2461 ccttagaaga tggttggctg gaaggttctc tgaaaggtcg taccggtatc tttccgtatc
     2521 gtttcgttaa actgtgcccg gcggccgcat aataaaccga tacaattaaa ggctcctttt
     2581 ggagcctttt ttttggagat tttcaacgtg aaaaaattat tattcgcaat tcctttagtt
     2641 gttcctttct attctcactc cgctgaaact gttgaaagtt gtttagcaaa accccataca
     2701 gaaaattcat ttactaacgt ctggaaagac gacaaaactt tagatcgtta cgctaactat
     2761 gagggctgtc tgtggaatgc tacaggcgtt gtagtttgta ctggtgacga aactcagtgt
     2821 ctagctagag tggcggtggc tctggttccg gtgattttga ttatgaaaag atggcaaacg
     2881 ctaataaggg ggctatgacc gaaaatgccg atgaaaacgc gctacagtct gacgctaaag
     2941 gcaaacttga ttctgtcgct actgattacg gtgctgctat cgatggtttc attggtgacg
     3001 tttccggcct tgctaatggt aatggtgcta ctggtgattt tgctggctct aattcccaaa
     3061 tggctcaagt cggtgacggt gataattcac ctttaatgaa taatttccgt caatatttac
     3121 cttccctccc tcaatcggtt gaatgtcgcc cttttgtctt tggcgctggt aaaccatatg
     3181 aattttctat tgattgtgac aaaataaact tattccgtgg tgtctttgcg tttcttttat
     3241 atgttgccac ctttatgtat gtattttcta cgtttgctaa catactgcgt aataaggagt
     3301 cttaatcatg ccagttcttt tggctagcgc cgccctatac cttgtctgcc tccccgcgtt
     3361 gcgtcgcggt gcatggagcc gggccacctc gacctgaatg gaagccggcg gcacctcgct
     3421 aacggattca ccactccaag aattggagcc aatcaattct tgcggagaac tgtgaatgcg
     3481 caaaccaacc cttggcagaa catatccatc gcgtccgcca tctccagcag ccgcacgcgg
     3541 cgcatctcgg gcagcgttgg gtcctggcca cgggtgcgca tgatcgtgct cctgtcgttg
     3601 aggacccggc taggctggcg gggttgcctt actggttagc agaatgaatc accgatacgc
     3661 gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg
     3721 gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt
     3781 atgttccgga tctgcatcgc aggatgctgc tggctaccct gtggaacacc tacatctgta
     3841 ttaacgaagc gctggcattg accctgagtg atttttctct ggtcccgccg catccatacc
     3901 gccagttgtt taccctcaca acgttccagt aaccgggcat gttcatcatc agtaacccgt
     3961 atcgtgagca tcctctctcg tttcatcggt atcattaccc ccatgaacag aaatccccct
     4021 tacacggagg catcagtgac caaacaggaa aaaaccgccc ttaacatggc ccgctttatc
     4081 agaagccaga cattaacgct tctggagaaa ctcaacgagc tggacgcgga tgaacaggca
     4141 gacatctgtg aatcgcttca cgaccacgct gatgagcttt accgcaggat ccgcgaaatt
     4201 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     4261 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     4321 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     4381 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     4441 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     4501 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     4561 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     4621 gccgcgctta atgcgccgct acagggcgcg tcgcggatcc tgcctcgcgc gtttcggtga
     4681 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     4741 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     4801 cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca
     4861 tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta
     4921 aggagaaaat accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4981 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5041 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5101 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5161 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccagggt
     5221 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     5281 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5341 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5401 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5461 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5521 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     5581 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5641 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5701 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5761 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5821 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5881 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5941 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     6001 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     6061 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     6121 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     6181 gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     6241 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccata ttgtgcaaaa
     6301 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6361 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6421 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6481 gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga actttaaaag
     6541 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6601 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6661 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6721 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6781 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6841 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     6901 tgacattaac ct
//
