LOCUS       sequence-192403-        6921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            ASAP2-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6921)
  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 6921)
  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_ASAP2-1/1
FEATURES             Location/Qualifiers
     source          1..6921
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     12..30
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        53..130
                     /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             131..1213
                     /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(150..169)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1456..1461,1462..1477,1478..1484)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1492..1508
                     /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     1497..1519
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1593..1610
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1626..2279
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2237..2259
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2286..2309
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2322..2339
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2571..2618
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4211..4666)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4348..4369)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4560..4579
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4708..4730)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4816..4956
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4871..4890
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4971..4988)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5142..5729)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5900..6691,6692..6760))
                     /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     6523..6542
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6761..6865)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 taacctataa aaataggcgt atcacgaggc cctttcgtct tcaagaattc ccgacaccat
       61 cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
      121 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
      181 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
      241 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
      301 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
      361 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
      421 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
      481 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
      541 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
      601 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
      661 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
      721 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
      781 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
      841 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
      901 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
      961 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     1021 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     1081 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     1141 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     1201 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcacaattct
     1261 catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc
     1321 atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa
     1381 ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt ctggcaaata
     1441 ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga attgtgagcg
     1501 gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga gcacttcacc
     1561 aacaaggacc atagattatg aaaatcgaag aacaccatca ccatcaccat tccagcggta
     1621 agcttatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa cccactcgac
     1681 ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc gatgaaggtg
     1741 ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt ccttattata
     1801 ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata gctgacaagc
     1861 acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt gaaggagcgg
     1921 ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt gaaactctca
     1981 aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat cgtttatgtc
     2041 ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg tatgacgctc
     2101 ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa ttagtttgtt
     2161 ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc agcaagtata
     2221 tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat cctccaaaat
     2281 cggatctgga agttctgttc caggggcccc tgtccagcgg tctggttccg cgtggttccg
     2341 gtaccgcggc ccagccggcc atggcccagg gcgcgctgaa acgtgtcaaa gccctgtaca
     2401 actgtgttgc cgataatcca gatgaactga ccttctccga aggtgatgtc attatcgtcg
     2461 atggtgaaga agatcaggaa tggtggattg gtcatattga tggtgatccg ggtcgtaaag
     2521 gtgcatttcc ggtttccttt gtccatttca ttgccgcggc cgcataataa accgatacaa
     2581 ttaaaggctc cttttggagc cttttttttg gagattttca acgtgaaaaa attattattc
     2641 gcaattcctt tagttgttcc tttctattct cactccgctg aaactgttga aagttgttta
     2701 gcaaaacccc atacagaaaa ttcatttact aacgtctgga aagacgacaa aactttagat
     2761 cgttacgcta actatgaggg ctgtctgtgg aatgctacag gcgttgtagt ttgtactggt
     2821 gacgaaactc agtgtctagc tagagtggcg gtggctctgg ttccggtgat tttgattatg
     2881 aaaagatggc aaacgctaat aagggggcta tgaccgaaaa tgccgatgaa aacgcgctac
     2941 agtctgacgc taaaggcaaa cttgattctg tcgctactga ttacggtgct gctatcgatg
     3001 gtttcattgg tgacgtttcc ggccttgcta atggtaatgg tgctactggt gattttgctg
     3061 gctctaattc ccaaatggct caagtcggtg acggtgataa ttcaccttta atgaataatt
     3121 tccgtcaata tttaccttcc ctccctcaat cggttgaatg tcgccctttt gtctttggcg
     3181 ctggtaaacc atatgaattt tctattgatt gtgacaaaat aaacttattc cgtggtgtct
     3241 ttgcgtttct tttatatgtt gccaccttta tgtatgtatt ttctacgttt gctaacatac
     3301 tgcgtaataa ggagtcttaa tcatgccagt tcttttggct agcgccgccc tataccttgt
     3361 ctgcctcccc gcgttgcgtc gcggtgcatg gagccgggcc acctcgacct gaatggaagc
     3421 cggcggcacc tcgctaacgg attcaccact ccaagaattg gagccaatca attcttgcgg
     3481 agaactgtga atgcgcaaac caacccttgg cagaacatat ccatcgcgtc cgccatctcc
     3541 agcagccgca cgcggcgcat ctcgggcagc gttgggtcct ggccacgggt gcgcatgatc
     3601 gtgctcctgt cgttgaggac ccggctaggc tggcggggtt gccttactgg ttagcagaat
     3661 gaatcaccga tacgcgagcg aacgtgaagc gactgctgct gcaaaacgtc tgcgacctga
     3721 gcaacaacat gaatggtctt cggtttccgt gtttcgtaaa gtctggaaac gcggaagtca
     3781 gcgccctgca ccattatgtt ccggatctgc atcgcaggat gctgctggct accctgtgga
     3841 acacctacat ctgtattaac gaagcgctgg cattgaccct gagtgatttt tctctggtcc
     3901 cgccgcatcc ataccgccag ttgtttaccc tcacaacgtt ccagtaaccg ggcatgttca
     3961 tcatcagtaa cccgtatcgt gagcatcctc tctcgtttca tcggtatcat tacccccatg
     4021 aacagaaatc ccccttacac ggaggcatca gtgaccaaac aggaaaaaac cgcccttaac
     4081 atggcccgct ttatcagaag ccagacatta acgcttctgg agaaactcaa cgagctggac
     4141 gcggatgaac aggcagacat ctgtgaatcg cttcacgacc acgctgatga gctttaccgc
     4201 aggatccgcg aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     4261 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     4321 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     4381 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     4441 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     4501 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     4561 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     4621 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcgcg gatcctgcct
     4681 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     4741 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     4801 tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtatactgg
     4861 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata
     4921 ccgcacagat gcgtaaggag aaaataccgc atcaggcgct cttccgcttc ctcgctcact
     4981 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5041 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5101 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5161 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5221 taaagatacc agggtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5281 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     5341 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     5401 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5461 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5521 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5581 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5641 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5701 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5761 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5821 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5881 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5941 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     6001 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6061 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     6121 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6181 cagttaatag tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt
     6241 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6301 ccatattgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6361 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     6421 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     6481 gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg ggataatacc gcgccacata
     6541 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     6601 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6661 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6721 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6781 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6841 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     6901 aaaccattat tatcatgaca t
//
