LOCUS       sequence-192600-        6897 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            PLCG1-1/1.
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_PLCG1-1/1
FEATURES             Location/Qualifiers
     source          1..6897
                     /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      2541..2588
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4181..4636)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4318..4339)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4530..4549
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4678..4700)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4786..4926
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4841..4860
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4941..4958)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5112..5699)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5870..6661,6662..6730))
                     /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     6493..6512
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6731..6835)
                     /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 ggcctgtgca gttaaagccc tgtttgatta caaagcccag cgtgaagatg
     2401 aactgacctt tatcaaatcc gccattatcc agaacgtcga aaaacaagaa ggtggttggt
     2461 ggcgtggtga ttatggtggt aaaaaacagc tgtggtttcc gtccaactac gtcgaagaaa
     2521 tggtcgcggc cgcataataa accgatacaa ttaaaggctc cttttggagc cttttttttg
     2581 gagattttca acgtgaaaaa attattattc gcaattcctt tagttgttcc tttctattct
     2641 cactccgctg aaactgttga aagttgttta gcaaaacccc atacagaaaa ttcatttact
     2701 aacgtctgga aagacgacaa aactttagat cgttacgcta actatgaggg ctgtctgtgg
     2761 aatgctacag gcgttgtagt ttgtactggt gacgaaactc agtgtctagc tagagtggcg
     2821 gtggctctgg ttccggtgat tttgattatg aaaagatggc aaacgctaat aagggggcta
     2881 tgaccgaaaa tgccgatgaa aacgcgctac agtctgacgc taaaggcaaa cttgattctg
     2941 tcgctactga ttacggtgct gctatcgatg gtttcattgg tgacgtttcc ggccttgcta
     3001 atggtaatgg tgctactggt gattttgctg gctctaattc ccaaatggct caagtcggtg
     3061 acggtgataa ttcaccttta atgaataatt tccgtcaata tttaccttcc ctccctcaat
     3121 cggttgaatg tcgccctttt gtctttggcg ctggtaaacc atatgaattt tctattgatt
     3181 gtgacaaaat aaacttattc cgtggtgtct ttgcgtttct tttatatgtt gccaccttta
     3241 tgtatgtatt ttctacgttt gctaacatac tgcgtaataa ggagtcttaa tcatgccagt
     3301 tcttttggct agcgccgccc tataccttgt ctgcctcccc gcgttgcgtc gcggtgcatg
     3361 gagccgggcc acctcgacct gaatggaagc cggcggcacc tcgctaacgg attcaccact
     3421 ccaagaattg gagccaatca attcttgcgg agaactgtga atgcgcaaac caacccttgg
     3481 cagaacatat ccatcgcgtc cgccatctcc agcagccgca cgcggcgcat ctcgggcagc
     3541 gttgggtcct ggccacgggt gcgcatgatc gtgctcctgt cgttgaggac ccggctaggc
     3601 tggcggggtt gccttactgg ttagcagaat gaatcaccga tacgcgagcg aacgtgaagc
     3661 gactgctgct gcaaaacgtc tgcgacctga gcaacaacat gaatggtctt cggtttccgt
     3721 gtttcgtaaa gtctggaaac gcggaagtca gcgccctgca ccattatgtt ccggatctgc
     3781 atcgcaggat gctgctggct accctgtgga acacctacat ctgtattaac gaagcgctgg
     3841 cattgaccct gagtgatttt tctctggtcc cgccgcatcc ataccgccag ttgtttaccc
     3901 tcacaacgtt ccagtaaccg ggcatgttca tcatcagtaa cccgtatcgt gagcatcctc
     3961 tctcgtttca tcggtatcat tacccccatg aacagaaatc ccccttacac ggaggcatca
     4021 gtgaccaaac aggaaaaaac cgcccttaac atggcccgct ttatcagaag ccagacatta
     4081 acgcttctgg agaaactcaa cgagctggac gcggatgaac aggcagacat ctgtgaatcg
     4141 cttcacgacc acgctgatga gctttaccgc aggatccgcg aaattgtaag cgttaatatt
     4201 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     4261 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     4321 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     4381 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     4441 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     4501 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     4561 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     4621 ccgctacagg gcgcgtcgcg gatcctgcct cgcgcgtttc ggtgatgacg gtgaaaacct
     4681 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     4741 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca
     4801 gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga gcagattgta
     4861 ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     4921 atcaggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4981 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     5041 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     5101 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5161 agtcagaggt ggcgaaaccc gacaggacta taaagatacc agggtttccc cctggaagct
     5221 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5281 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5341 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5401 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5461 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5521 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     5581 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5641 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5701 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5761 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     5821 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     5881 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     5941 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6001 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6061 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6121 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6181 ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6241 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6301 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6361 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     6421 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6481 cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6541 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6601 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6661 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6721 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     6781 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     6841 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacct
//
