LOCUS       sequence-192277-        6963 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            PIK3R2-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6963)
  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 6963)
  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_PIK3R2-1/1
FEATURES             Location/Qualifiers
     source          1..6963
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        40..117
                     /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             118..1200
                     /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(137..156)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1443..1448,1449..1464,1465..1471)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1479..1495
                     /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     1484..1506
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1580..1597
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1613..2266
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2224..2246
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2273..2296
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2309..2326
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2600..2647
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4240..4695)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4377..4398)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4589..4608
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4737..4759)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4845..4985
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4900..4919
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5000..5017)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5171..5758)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5929..6720,6721..6789))
                     /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     6552..6571
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6790..6894)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     join(6962..6963,1..17)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 taggcgtatc acgaggccct ttcgtcttca agaattcccg acaccatcga atggtgcaaa
       61 acctttcgcg gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatgtg
      121 aaaccagtaa cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc
      181 cgcgtggtga accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg
      241 atggcggagc tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg
      301 ttgctgattg gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg
      361 gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga
      421 agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg
      481 ctgatcatta actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact
      541 aatgttccgg cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttc
      601 tcccatgaag acggtacgcg actgggcgtg gagcatctgg tcgcattggg tcaccagcaa
      661 atcgcgctgt tagcgggccc attaagttct gtctcggcgc gtctgcgtct ggctggctgg
      721 cataaatatc tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg cgactggagt
      781 gccatgtccg gttttcaaca aaccatgcaa atgctgaatg agggcatcgt tcccactgcg
      841 atgctggttg ccaacgatca gatggcgctg ggcgcaatgc gcgccattac cgagtccggg
      901 ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg ataccgaaga cagctcatgt
      961 tatatcccgc cgttaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg
     1021 gaccgcttgc tgcaactctc tcagggccag gcggtgaagg gcaatcagct gttgcccgtc
     1081 tcactggtga aaagaaaaac caccctggcg cccaatacgc aaaccgcctc tccccgcgcg
     1141 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     1201 gcgcaacgca attaatgtaa gttagctcac tcattaggca caattctcat gtttgacagc
     1261 ttatcatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
     1321 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
     1381 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
     1441 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
     1501 cacaggaaac agccagtccg tttaggtgtt ttcacgagca cttcaccaac aaggaccata
     1561 gattatgaaa atcgaagaac accatcacca tcaccattcc agcggtaagc ttatgtcccc
     1621 tatactaggt tattggaaaa ttaagggcct tgtgcaaccc actcgacttc ttttggaata
     1681 tcttgaagaa aaatatgaag agcatttgta tgagcgcgat gaaggtgata aatggcgaaa
     1741 caaaaagttt gaattgggtt tggagtttcc caatcttcct tattatattg atggtgatgt
     1801 taaattaaca cagtctatgg ccatcatacg ttatatagct gacaagcaca acatgttggg
     1861 tggttgtcca aaagagcgtg cagagatttc aatgcttgaa ggagcggttt tggatattag
     1921 atacggtgtt tcgagaattg catatagtaa agactttgaa actctcaaag ttgattttct
     1981 tagcaagcta cctgaaatgc tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt
     2041 aaatggtgat catgtaaccc atcctgactt catgttgtat gacgctcttg atgttgtttt
     2101 atacatggac ccaatgtgcc tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat
     2161 tgaagctatc ccacaaattg ataagtactt gaaatccagc aagtatatag catggccttt
     2221 gcagggctgg caagccacgt ttggtggtgg cgaccatcct ccaaaatcgg atctggaagt
     2281 tctgttccag gggcccctgt ccagcggtct ggttccgcgt ggttccggta ccgcggccca
     2341 gccggccatg gcccagggcg cgctgtttca gtatcgtgcc ctgtatccat ttcgtcgtga
     2401 acgtccggaa gatttagaac tgctgccggg tgatgtttta gttgtttctc gtgcagcact
     2461 gcaagcatta ggtgttgcag aaggtggtga acgttgtcca caatccgttg gttggatgcc
     2521 aggtctgaat gaacgtaccc gtcagcgtgg tgattttccc ggtacttacg ttgaatttct
     2581 gggtgcggcc gcataataaa ccgatacaat taaaggctcc ttttggagcc ttttttttgg
     2641 agattttcaa cgtgaaaaaa ttattattcg caattccttt agttgttcct ttctattctc
     2701 actccgctga aactgttgaa agttgtttag caaaacccca tacagaaaat tcatttacta
     2761 acgtctggaa agacgacaaa actttagatc gttacgctaa ctatgagggc tgtctgtgga
     2821 atgctacagg cgttgtagtt tgtactggtg acgaaactca gtgtctagct agagtggcgg
     2881 tggctctggt tccggtgatt ttgattatga aaagatggca aacgctaata agggggctat
     2941 gaccgaaaat gccgatgaaa acgcgctaca gtctgacgct aaaggcaaac ttgattctgt
     3001 cgctactgat tacggtgctg ctatcgatgg tttcattggt gacgtttccg gccttgctaa
     3061 tggtaatggt gctactggtg attttgctgg ctctaattcc caaatggctc aagtcggtga
     3121 cggtgataat tcacctttaa tgaataattt ccgtcaatat ttaccttccc tccctcaatc
     3181 ggttgaatgt cgcccttttg tctttggcgc tggtaaacca tatgaatttt ctattgattg
     3241 tgacaaaata aacttattcc gtggtgtctt tgcgtttctt ttatatgttg ccacctttat
     3301 gtatgtattt tctacgtttg ctaacatact gcgtaataag gagtcttaat catgccagtt
     3361 cttttggcta gcgccgccct ataccttgtc tgcctccccg cgttgcgtcg cggtgcatgg
     3421 agccgggcca cctcgacctg aatggaagcc ggcggcacct cgctaacgga ttcaccactc
     3481 caagaattgg agccaatcaa ttcttgcgga gaactgtgaa tgcgcaaacc aacccttggc
     3541 agaacatatc catcgcgtcc gccatctcca gcagccgcac gcggcgcatc tcgggcagcg
     3601 ttgggtcctg gccacgggtg cgcatgatcg tgctcctgtc gttgaggacc cggctaggct
     3661 ggcggggttg ccttactggt tagcagaatg aatcaccgat acgcgagcga acgtgaagcg
     3721 actgctgctg caaaacgtct gcgacctgag caacaacatg aatggtcttc ggtttccgtg
     3781 tttcgtaaag tctggaaacg cggaagtcag cgccctgcac cattatgttc cggatctgca
     3841 tcgcaggatg ctgctggcta ccctgtggaa cacctacatc tgtattaacg aagcgctggc
     3901 attgaccctg agtgattttt ctctggtccc gccgcatcca taccgccagt tgtttaccct
     3961 cacaacgttc cagtaaccgg gcatgttcat catcagtaac ccgtatcgtg agcatcctct
     4021 ctcgtttcat cggtatcatt acccccatga acagaaatcc cccttacacg gaggcatcag
     4081 tgaccaaaca ggaaaaaacc gcccttaaca tggcccgctt tatcagaagc cagacattaa
     4141 cgcttctgga gaaactcaac gagctggacg cggatgaaca ggcagacatc tgtgaatcgc
     4201 ttcacgacca cgctgatgag ctttaccgca ggatccgcga aattgtaagc gttaatattt
     4261 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     4321 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     4381 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     4441 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     4501 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     4561 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     4621 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     4681 cgctacaggg cgcgtcgcgg atcctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc
     4741 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     4801 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag
     4861 tcacgtagcg atagcggagt gtatactggc ttaactatgc ggcatcagag cagattgtac
     4921 tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     4981 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5041 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5101 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5161 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5221 gtcagaggtg gcgaaacccg acaggactat aaagatacca gggtttcccc ctggaagctc
     5281 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     5341 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5401 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     5461 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     5521 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     5581 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     5641 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     5701 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     5761 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5821 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5881 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5941 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     6001 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     6061 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     6121 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     6181 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     6241 tgctgcaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     6301 ccaacgatca aggcgagtta catgatcccc catattgtgc aaaaaagcgg ttagctcctt
     6361 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     6421 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     6481 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     6541 gtcaacacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     6601 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     6661 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     6721 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     6781 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6841 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6901 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     6961 aaa
//
