LOCUS       sequence-191790-        6930 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            TJP1-1/1-SV1/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6930)
  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 6930)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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_TJP1-1/1-SV1/2
FEATURES             Location/Qualifiers
     source          1..6930
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..39
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        62..139
                     /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             140..1222
                     /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(159..178)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1465..1470,1471..1486,1487..1493)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1501..1517
                     /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     1506..1528
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1602..1619
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1635..2288
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2246..2268
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2295..2318
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2331..2348
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2589..2636
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4229..4684)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4366..4387)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4578..4597
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4726..4748)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4834..4974
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4889..4908
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4989..5006)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5160..5747)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5918..6709,6710..6778))
                     /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     6541..6560
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6779..6883)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctt caagaattcc
       61 cgacaccatc gaatggtgca aaacctttcg cggtatggca tgatagcgcc cggaagagag
      121 tcaattcagg gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg
      181 tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac
      241 gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca
      301 acaactggcg ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca
      361 cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt
      421 ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct
      481 tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat
      541 tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac
      601 acccatcaac agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct
      661 ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc
      721 gcgtctgcgt ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc
      781 ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa
      841 tgagggcatc gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat
      901 gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga
      961 cgataccgaa gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg
     1021 cctgctgggg caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa
     1081 gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac
     1141 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
     1201 ccgactggaa agcgggcagt gagcgcaacg caattaatgt aagttagctc actcattagg
     1261 cacaattctc atgtttgaca gcttatcatc gactgcacgg tgcaccaatg cttctggcgt
     1321 caggcagcca tcggaagctg tggtatggct gtgcaggtcg taaatcactg cataattcgt
     1381 gtcgctcaag gcgcactccc gttctggata atgttttttg cgccgacatc ataacggttc
     1441 tggcaaatat tctgaaatga gctgttgaca attaatcatc ggctcgtata atgtgtggaa
     1501 ttgtgagcgg ataacaattt cacacaggaa acagccagtc cgtttaggtg ttttcacgag
     1561 cacttcacca acaaggacca tagattatga aaatcgaaga acaccatcac catcaccatt
     1621 ccagcggtaa gcttatgtcc cctatactag gttattggaa aattaagggc cttgtgcaac
     1681 ccactcgact tcttttggaa tatcttgaag aaaaatatga agagcatttg tatgagcgcg
     1741 atgaaggtga taaatggcga aacaaaaagt ttgaattggg tttggagttt cccaatcttc
     1801 cttattatat tgatggtgat gttaaattaa cacagtctat ggccatcata cgttatatag
     1861 ctgacaagca caacatgttg ggtggttgtc caaaagagcg tgcagagatt tcaatgcttg
     1921 aaggagcggt tttggatatt agatacggtg tttcgagaat tgcatatagt aaagactttg
     1981 aaactctcaa agttgatttt cttagcaagc tacctgaaat gctgaaaatg ttcgaagatc
     2041 gtttatgtca taaaacatat ttaaatggtg atcatgtaac ccatcctgac ttcatgttgt
     2101 atgacgctct tgatgttgtt ttatacatgg acccaatgtg cctggatgcg ttcccaaaat
     2161 tagtttgttt taaaaaacgt attgaagcta tcccacaaat tgataagtac ttgaaatcca
     2221 gcaagtatat agcatggcct ttgcagggct ggcaagccac gtttggtggt ggcgaccatc
     2281 ctccaaaatc ggatctggaa gttctgttcc aggggcccct gtccagcggt ctggttccgc
     2341 gtggttccgg taccgcggcc cagccggccg gcgatagttt ctacatccgt acccatttcg
     2401 aatatgaaaa agaatctccg tatggtctgt cgttcaataa aggcgaagtg ttccgtgtgg
     2461 ttgataccct gtataacggc aaactgggta gctggctggc gattcgtatc ggcaaaaatc
     2521 ataaagaagt cgaacgtggt attatcccga acaaaaatcg cgccgaacag ctggcggccg
     2581 cataataaac cgatacaatt aaaggctcct tttggagcct tttttttgga gattttcaac
     2641 gtgaaaaaat tattattcgc aattccttta gttgttcctt tctattctca ctccgctgaa
     2701 actgttgaaa gttgtttagc aaaaccccat acagaaaatt catttactaa cgtctggaaa
     2761 gacgacaaaa ctttagatcg ttacgctaac tatgagggct gtctgtggaa tgctacaggc
     2821 gttgtagttt gtactggtga cgaaactcag tgtctagcta gagtggcggt ggctctggtt
     2881 ccggtgattt tgattatgaa aagatggcaa acgctaataa gggggctatg accgaaaatg
     2941 ccgatgaaaa cgcgctacag tctgacgcta aaggcaaact tgattctgtc gctactgatt
     3001 acggtgctgc tatcgatggt ttcattggtg acgtttccgg ccttgctaat ggtaatggtg
     3061 ctactggtga ttttgctggc tctaattccc aaatggctca agtcggtgac ggtgataatt
     3121 cacctttaat gaataatttc cgtcaatatt taccttccct ccctcaatcg gttgaatgtc
     3181 gcccttttgt ctttggcgct ggtaaaccat atgaattttc tattgattgt gacaaaataa
     3241 acttattccg tggtgtcttt gcgtttcttt tatatgttgc cacctttatg tatgtatttt
     3301 ctacgtttgc taacatactg cgtaataagg agtcttaatc atgccagttc ttttggctag
     3361 cgccgcccta taccttgtct gcctccccgc gttgcgtcgc ggtgcatgga gccgggccac
     3421 ctcgacctga atggaagccg gcggcacctc gctaacggat tcaccactcc aagaattgga
     3481 gccaatcaat tcttgcggag aactgtgaat gcgcaaacca acccttggca gaacatatcc
     3541 atcgcgtccg ccatctccag cagccgcacg cggcgcatct cgggcagcgt tgggtcctgg
     3601 ccacgggtgc gcatgatcgt gctcctgtcg ttgaggaccc ggctaggctg gcggggttgc
     3661 cttactggtt agcagaatga atcaccgata cgcgagcgaa cgtgaagcga ctgctgctgc
     3721 aaaacgtctg cgacctgagc aacaacatga atggtcttcg gtttccgtgt ttcgtaaagt
     3781 ctggaaacgc ggaagtcagc gccctgcacc attatgttcc ggatctgcat cgcaggatgc
     3841 tgctggctac cctgtggaac acctacatct gtattaacga agcgctggca ttgaccctga
     3901 gtgatttttc tctggtcccg ccgcatccat accgccagtt gtttaccctc acaacgttcc
     3961 agtaaccggg catgttcatc atcagtaacc cgtatcgtga gcatcctctc tcgtttcatc
     4021 ggtatcatta cccccatgaa cagaaatccc ccttacacgg aggcatcagt gaccaaacag
     4081 gaaaaaaccg cccttaacat ggcccgcttt atcagaagcc agacattaac gcttctggag
     4141 aaactcaacg agctggacgc ggatgaacag gcagacatct gtgaatcgct tcacgaccac
     4201 gctgatgagc tttaccgcag gatccgcgaa attgtaagcg ttaatatttt gttaaaattc
     4261 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
     4321 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
     4381 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
     4441 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa
     4501 gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg
     4561 aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt
     4621 gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc
     4681 gcgtcgcgga tcctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     4741 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     4801 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     4861 tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac
     4921 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct
     4981 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5041 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5101 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5161 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     5221 cgaaacccga caggactata aagataccag ggtttccccc tggaagctcc ctcgtgcgct
     5281 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     5341 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     5401 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     5461 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     5521 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     5581 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     5641 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     5701 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     5761 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     5821 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     5881 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     5941 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     6001 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     6061 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     6121 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     6181 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca
     6241 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     6301 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     6361 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     6421 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     6481 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaacacggg
     6541 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     6601 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     6661 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     6721 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     6781 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     6841 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     6901 tgccacctga cgtctaagaa accattatta
//
