LOCUS       sequence-192258-        6957 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            LASP1-1/1-SV2/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6957)
  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 6957)
  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_LASP1-1/1-SV2/2
FEATURES             Location/Qualifiers
     source          1..6957
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     24..42
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        65..142
                     /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             143..1225
                     /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(162..181)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1468..1473,1474..1489,1490..1496)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1504..1520
                     /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     1509..1531
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1605..1622
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1638..2291
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2249..2271
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2298..2321
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2334..2351
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2571..2600
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      2619..2666
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4259..4714)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4396..4417)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4608..4627
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4756..4778)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4864..5004
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4919..4938
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5019..5036)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5190..5777)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5948..6739,6740..6808))
                     /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     6571..6590
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6809..6913)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt cttcaagaat
       61 tcccgacacc atcgaatggt gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga
      121 gagtcaattc agggtggtga atgtgaaacc agtaacgtta tacgatgtcg cagagtatgc
      181 cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa
      241 aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc
      301 acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct
      361 gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag
      421 cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa
      481 tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc
      541 cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca
      601 gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg gcgtggagca
      661 tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttctgtctc
      721 ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa ttcagccgat
      781 agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca tgcaaatgct
      841 gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc
      901 aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg tagtgggata
      961 cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca aacaggattt
     1021 tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt
     1081 gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc tggcgcccaa
     1141 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     1201 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtaagttag ctcactcatt
     1261 aggcacaatt ctcatgtttg acagcttatc atcgactgca cggtgcacca atgcttctgg
     1321 cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca ctgcataatt
     1381 cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt ttgcgccgac atcataacgg
     1441 ttctggcaaa tattctgaaa tgagctgttg acaattaatc atcggctcgt ataatgtgtg
     1501 gaattgtgag cggataacaa tttcacacag gaaacagcca gtccgtttag gtgttttcac
     1561 gagcacttca ccaacaagga ccatagatta tgaaaatcga agaacaccat caccatcacc
     1621 attccagcgg taagcttatg tcccctatac taggttattg gaaaattaag ggccttgtgc
     1681 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
     1741 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
     1801 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
     1861 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
     1921 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
     1981 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
     2041 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
     2101 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
     2161 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
     2221 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
     2281 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgtccagc ggtctggttc
     2341 cgcgtggttc cggtaccgcg gcccagccgg ccatggccca gggcgcgctg aaacgttatc
     2401 gtgccgtcta tgattactct gccgccgatg aagatgaagt ctcctttcag gatggtgata
     2461 ccattgttaa cgttcagcag attgatgatg gttggatgta cggtactgtt gaacgtaccg
     2521 gtgatactgg tatgctgcca gccaactatg tcgaagccat tgcggccgca gaacaaaaac
     2581 tcatctcaga agaggatctg aatggggccg cataataaac cgatacaatt aaaggctcct
     2641 tttggagcct tttttttgga gattttcaac gtgaaaaaat tattattcgc aattccttta
     2701 gttgttcctt tctattctca ctccgctgaa actgttgaaa gttgtttagc aaaaccccat
     2761 acagaaaatt catttactaa cgtctggaaa gacgacaaaa ctttagatcg ttacgctaac
     2821 tatgagggct gtctgtggaa tgctacaggc gttgtagttt gtactggtga cgaaactcag
     2881 tgtctagcta gagtggcggt ggctctggtt ccggtgattt tgattatgaa aagatggcaa
     2941 acgctaataa gggggctatg accgaaaatg ccgatgaaaa cgcgctacag tctgacgcta
     3001 aaggcaaact tgattctgtc gctactgatt acggtgctgc tatcgatggt ttcattggtg
     3061 acgtttccgg ccttgctaat ggtaatggtg ctactggtga ttttgctggc tctaattccc
     3121 aaatggctca agtcggtgac ggtgataatt cacctttaat gaataatttc cgtcaatatt
     3181 taccttccct ccctcaatcg gttgaatgtc gcccttttgt ctttggcgct ggtaaaccat
     3241 atgaattttc tattgattgt gacaaaataa acttattccg tggtgtcttt gcgtttcttt
     3301 tatatgttgc cacctttatg tatgtatttt ctacgtttgc taacatactg cgtaataagg
     3361 agtcttaatc atgccagttc ttttggctag cgccgcccta taccttgtct gcctccccgc
     3421 gttgcgtcgc ggtgcatgga gccgggccac ctcgacctga atggaagccg gcggcacctc
     3481 gctaacggat tcaccactcc aagaattgga gccaatcaat tcttgcggag aactgtgaat
     3541 gcgcaaacca acccttggca gaacatatcc atcgcgtccg ccatctccag cagccgcacg
     3601 cggcgcatct cgggcagcgt tgggtcctgg ccacgggtgc gcatgatcgt gctcctgtcg
     3661 ttgaggaccc ggctaggctg gcggggttgc cttactggtt agcagaatga atcaccgata
     3721 cgcgagcgaa cgtgaagcga ctgctgctgc aaaacgtctg cgacctgagc aacaacatga
     3781 atggtcttcg gtttccgtgt ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc
     3841 attatgttcc ggatctgcat cgcaggatgc tgctggctac cctgtggaac acctacatct
     3901 gtattaacga agcgctggca ttgaccctga gtgatttttc tctggtcccg ccgcatccat
     3961 accgccagtt gtttaccctc acaacgttcc agtaaccggg catgttcatc atcagtaacc
     4021 cgtatcgtga gcatcctctc tcgtttcatc ggtatcatta cccccatgaa cagaaatccc
     4081 ccttacacgg aggcatcagt gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt
     4141 atcagaagcc agacattaac gcttctggag aaactcaacg agctggacgc ggatgaacag
     4201 gcagacatct gtgaatcgct tcacgaccac gctgatgagc tttaccgcag gatccgcgaa
     4261 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     4321 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     4381 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     4441 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     4501 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     4561 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     4621 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     4681 cccgccgcgc ttaatgcgcc gctacagggc gcgtcgcgga tcctgcctcg cgcgtttcgg
     4741 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta
     4801 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
     4861 gggcgcagcc atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg
     4921 gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc
     4981 gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5041 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5101 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5161 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5221 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5281 ggtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5341 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5401 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5461 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5521 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5581 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg
     5641 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5701 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5761 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5821 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5881 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     5941 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     6001 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     6061 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     6121 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6181 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     6241 tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6301 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atattgtgca
     6361 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6421 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6481 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6541 cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa
     6601 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6661 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6721 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6781 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6841 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     6901 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accatta
//
