LOCUS       sequence-191131-        6954 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            FYB-2/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6954)
  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 6954)
  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_FYB-2/2
FEATURES             Location/Qualifiers
     source          1..6954
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     19..37
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        60..137
                     /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             138..1220
                     /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(157..176)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1463..1468,1469..1484,1485..1491)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1499..1515
                     /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     1504..1526
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1600..1617
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1633..2286
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2244..2266
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2293..2316
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2329..2346
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2611..2658
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4251..4706)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4388..4409)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4600..4619
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4748..4770)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4856..4996
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4911..4930
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5011..5028)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5182..5769)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5940..6731,6732..6800))
                     /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     6563..6582
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6801..6905)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtcttca agaattcccg
       61 acaccatcga atggtgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc
      121 aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg
      181 tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc
      241 gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac
      301 aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg
      361 cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg
      421 tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc
      481 tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg
      541 ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac
      601 ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg gagcatctgg
      661 tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc
      721 gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg
      781 aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg
      841 agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc
      901 gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg
      961 ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc
     1021 tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg
     1081 gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg cccaatacgc
     1141 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     1201 gactggaaag cgggcagtga gcgcaacgca attaatgtaa gttagctcac tcattaggca
     1261 caattctcat gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca
     1321 ggcagccatc ggaagctgtg gtatggctgt gcaggtcgta aatcactgca taattcgtgt
     1381 cgctcaaggc gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg
     1441 gcaaatattc tgaaatgagc tgttgacaat taatcatcgg ctcgtataat gtgtggaatt
     1501 gtgagcggat aacaatttca cacaggaaac agccagtccg tttaggtgtt ttcacgagca
     1561 cttcaccaac aaggaccata gattatgaaa atcgaagaac accatcacca tcaccattcc
     1621 agcggtaagc ttatgtcccc tatactaggt tattggaaaa ttaagggcct tgtgcaaccc
     1681 actcgacttc ttttggaata tcttgaagaa aaatatgaag agcatttgta tgagcgcgat
     1741 gaaggtgata aatggcgaaa caaaaagttt gaattgggtt tggagtttcc caatcttcct
     1801 tattatattg atggtgatgt taaattaaca cagtctatgg ccatcatacg ttatatagct
     1861 gacaagcaca acatgttggg tggttgtcca aaagagcgtg cagagatttc aatgcttgaa
     1921 ggagcggttt tggatattag atacggtgtt tcgagaattg catatagtaa agactttgaa
     1981 actctcaaag ttgattttct tagcaagcta cctgaaatgc tgaaaatgtt cgaagatcgt
     2041 ttatgtcata aaacatattt aaatggtgat catgtaaccc atcctgactt catgttgtat
     2101 gacgctcttg atgttgtttt atacatggac ccaatgtgcc tggatgcgtt cccaaaatta
     2161 gtttgtttta aaaaacgtat tgaagctatc ccacaaattg ataagtactt gaaatccagc
     2221 aagtatatag catggccttt gcagggctgg caagccacgt ttggtggtgg cgaccatcct
     2281 ccaaaatcgg atctggaagt tctgttccag gggcccctgt ccagcggtct ggttccgcgt
     2341 ggttccggta ccgcggccca gccggccgaa aaagaagaaa aagacttccg taaaaaattc
     2401 aaatatgatg gcgaaatccg tgtcctgtac tcgacgaaag tgacgacctc aattaccagc
     2461 aaaaaatggg gcacgcgtga tctgcaggtg aaaccgggtg aatctctgga agttattcaa
     2521 accacggatg acaccaaagt cctgtgccgt aacgaagaag gcaaatatgg ttacgttctg
     2581 cgcagttatc tggcggcggc cgcataataa accgatacaa ttaaaggctc cttttggagc
     2641 cttttttttg gagattttca acgtgaaaaa attattattc gcaattcctt tagttgttcc
     2701 tttctattct cactccgctg aaactgttga aagttgttta gcaaaacccc atacagaaaa
     2761 ttcatttact aacgtctgga aagacgacaa aactttagat cgttacgcta actatgaggg
     2821 ctgtctgtgg aatgctacag gcgttgtagt ttgtactggt gacgaaactc agtgtctagc
     2881 tagagtggcg gtggctctgg ttccggtgat tttgattatg aaaagatggc aaacgctaat
     2941 aagggggcta tgaccgaaaa tgccgatgaa aacgcgctac agtctgacgc taaaggcaaa
     3001 cttgattctg tcgctactga ttacggtgct gctatcgatg gtttcattgg tgacgtttcc
     3061 ggccttgcta atggtaatgg tgctactggt gattttgctg gctctaattc ccaaatggct
     3121 caagtcggtg acggtgataa ttcaccttta atgaataatt tccgtcaata tttaccttcc
     3181 ctccctcaat cggttgaatg tcgccctttt gtctttggcg ctggtaaacc atatgaattt
     3241 tctattgatt gtgacaaaat aaacttattc cgtggtgtct ttgcgtttct tttatatgtt
     3301 gccaccttta tgtatgtatt ttctacgttt gctaacatac tgcgtaataa ggagtcttaa
     3361 tcatgccagt tcttttggct agcgccgccc tataccttgt ctgcctcccc gcgttgcgtc
     3421 gcggtgcatg gagccgggcc acctcgacct gaatggaagc cggcggcacc tcgctaacgg
     3481 attcaccact ccaagaattg gagccaatca attcttgcgg agaactgtga atgcgcaaac
     3541 caacccttgg cagaacatat ccatcgcgtc cgccatctcc agcagccgca cgcggcgcat
     3601 ctcgggcagc gttgggtcct ggccacgggt gcgcatgatc gtgctcctgt cgttgaggac
     3661 ccggctaggc tggcggggtt gccttactgg ttagcagaat gaatcaccga tacgcgagcg
     3721 aacgtgaagc gactgctgct gcaaaacgtc tgcgacctga gcaacaacat gaatggtctt
     3781 cggtttccgt gtttcgtaaa gtctggaaac gcggaagtca gcgccctgca ccattatgtt
     3841 ccggatctgc atcgcaggat gctgctggct accctgtgga acacctacat ctgtattaac
     3901 gaagcgctgg cattgaccct gagtgatttt tctctggtcc cgccgcatcc ataccgccag
     3961 ttgtttaccc tcacaacgtt ccagtaaccg ggcatgttca tcatcagtaa cccgtatcgt
     4021 gagcatcctc tctcgtttca tcggtatcat tacccccatg aacagaaatc ccccttacac
     4081 ggaggcatca gtgaccaaac aggaaaaaac cgcccttaac atggcccgct ttatcagaag
     4141 ccagacatta acgcttctgg agaaactcaa cgagctggac gcggatgaac aggcagacat
     4201 ctgtgaatcg cttcacgacc acgctgatga gctttaccgc aggatccgcg aaattgtaag
     4261 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     4321 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     4381 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     4441 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     4501 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     4561 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     4621 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     4681 gcttaatgcg ccgctacagg gcgcgtcgcg gatcctgcct cgcgcgtttc ggtgatgacg
     4741 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     4801 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag
     4861 ccatgaccca gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga
     4921 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     4981 aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5041 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5101 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5161 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5221 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc agggtttccc
     5281 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5341 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     5401 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5461 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5521 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     5581 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     5641 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     5701 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     5761 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     5821 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5881 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     5941 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6001 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6061 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6121 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6181 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6241 gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6301 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6361 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6421 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6481 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6541 tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     6601 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     6661 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     6721 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6781 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6841 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6901 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatc
//
