LOCUS       sequence-192229-        6936 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            RIMBP3-3/3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6936)
  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 6936)
  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_RIMBP3-3/3
FEATURES             Location/Qualifiers
     source          1..6936
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     13..31
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        54..131
                     /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             132..1214
                     /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(151..170)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1457..1462,1463..1478,1479..1485)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1493..1509
                     /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     1498..1520
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1594..1611
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1627..2280
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2238..2260
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2287..2310
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2323..2340
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2587..2634
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4227..4682)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4364..4385)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4576..4595
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4724..4746)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4832..4972
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4887..4906
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4987..5004)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5158..5745)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5916..6707,6708..6776))
                     /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     6539..6558
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6777..6881)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttaacctata aaaataggcg tatcacgagg ccctttcgtc ttcaagaatt cccgacacca
       61 tcgaatggtg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
      121 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
      181 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
      241 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
      301 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
      361 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
      421 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
      481 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
      541 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca
      601 acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat
      661 tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc
      721 gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg
      781 aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca
      841 tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca
      901 ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg
      961 aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg
     1021 ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc
     1081 agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg
     1141 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     1201 aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcacaattc
     1261 tcatgtttga cagcttatca tcgactgcac ggtgcaccaa tgcttctggc gtcaggcagc
     1321 catcggaagc tgtggtatgg ctgtgcaggt cgtaaatcac tgcataattc gtgtcgctca
     1381 aggcgcactc ccgttctgga taatgttttt tgcgccgaca tcataacggt tctggcaaat
     1441 attctgaaat gagctgttga caattaatca tcggctcgta taatgtgtgg aattgtgagc
     1501 ggataacaat ttcacacagg aaacagccag tccgtttagg tgttttcacg agcacttcac
     1561 caacaaggac catagattat gaaaatcgaa gaacaccatc accatcacca ttccagcggt
     1621 aagcttatgt cccctatact aggttattgg aaaattaagg gccttgtgca acccactcga
     1681 cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg cgatgaaggt
     1741 gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct tccttattat
     1801 attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat agctgacaag
     1861 cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct tgaaggagcg
     1921 gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt tgaaactctc
     1981 aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga tcgtttatgt
     2041 cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt gtatgacgct
     2101 cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa attagtttgt
     2161 tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc cagcaagtat
     2221 atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca tcctccaaaa
     2281 tcggatctgg aagttctgtt ccaggggccc ctgtccagcg gtctggttcc gcgtggttcc
     2341 ggtaccgcgg cccagccggc catggcccag ggcgcgctga aaatcatgat tgccgccctg
     2401 gattatgatc caggtgatgg tcaaatgggt ggtcaaggta aaggtcgtct ggcattacgt
     2461 gcaggtgatg tcgttatggt ttatggtccg atggatgatc agggtttcta ctacggtgaa
     2521 ctgggtggtc atcgtggttt agttccagca catctgctgg atcacatgtc tgcggccgca
     2581 taataaaccg atacaattaa aggctccttt tggagccttt tttttggaga ttttcaacgt
     2641 gaaaaaatta ttattcgcaa ttcctttagt tgttcctttc tattctcact ccgctgaaac
     2701 tgttgaaagt tgtttagcaa aaccccatac agaaaattca tttactaacg tctggaaaga
     2761 cgacaaaact ttagatcgtt acgctaacta tgagggctgt ctgtggaatg ctacaggcgt
     2821 tgtagtttgt actggtgacg aaactcagtg tctagctaga gtggcggtgg ctctggttcc
     2881 ggtgattttg attatgaaaa gatggcaaac gctaataagg gggctatgac cgaaaatgcc
     2941 gatgaaaacg cgctacagtc tgacgctaaa ggcaaacttg attctgtcgc tactgattac
     3001 ggtgctgcta tcgatggttt cattggtgac gtttccggcc ttgctaatgg taatggtgct
     3061 actggtgatt ttgctggctc taattcccaa atggctcaag tcggtgacgg tgataattca
     3121 cctttaatga ataatttccg tcaatattta ccttccctcc ctcaatcggt tgaatgtcgc
     3181 ccttttgtct ttggcgctgg taaaccatat gaattttcta ttgattgtga caaaataaac
     3241 ttattccgtg gtgtctttgc gtttctttta tatgttgcca cctttatgta tgtattttct
     3301 acgtttgcta acatactgcg taataaggag tcttaatcat gccagttctt ttggctagcg
     3361 ccgccctata ccttgtctgc ctccccgcgt tgcgtcgcgg tgcatggagc cgggccacct
     3421 cgacctgaat ggaagccggc ggcacctcgc taacggattc accactccaa gaattggagc
     3481 caatcaattc ttgcggagaa ctgtgaatgc gcaaaccaac ccttggcaga acatatccat
     3541 cgcgtccgcc atctccagca gccgcacgcg gcgcatctcg ggcagcgttg ggtcctggcc
     3601 acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc ggggttgcct
     3661 tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact gctgctgcaa
     3721 aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt cgtaaagtct
     3781 ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg caggatgctg
     3841 ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt gaccctgagt
     3901 gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac aacgttccag
     3961 taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc gtttcatcgg
     4021 tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga ccaaacagga
     4081 aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc ttctggagaa
     4141 actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc acgaccacgc
     4201 tgatgagctt taccgcagga tccgcgaaat tgtaagcgtt aatattttgt taaaattcgc
     4261 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     4321 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     4381 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     4441 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     4501 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     4561 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     4621 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     4681 gtcgcggatc ctgcctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
     4741 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
     4801 gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca cgtagcgata
     4861 gcggagtgta tactggctta actatgcggc atcagagcag attgtactga gagtgcacca
     4921 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc
     4981 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     5041 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     5101 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     5161 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     5221 aaacccgaca ggactataaa gataccaggg tttccccctg gaagctccct cgtgcgctct
     5281 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5341 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5401 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5461 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5521 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5581 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5641 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5701 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5761 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     5821 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     5881 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     5941 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6001 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6061 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6121 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6181 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tgcaggcatc
     6241 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6301 cgagttacat gatcccccat attgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6361 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6421 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6481 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aacacgggat
     6541 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6601 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6661 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6721 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6781 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     6841 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     6901 ccacctgacg tctaagaaac cattattatc atgaca
//
