LOCUS       sequence-191811-        6918 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            PACSIN2-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6918)
  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 6918)
  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_PACSIN2-1/1
FEATURES             Location/Qualifiers
     source          1..6918
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     25..43
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        66..143
                     /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             144..1226
                     /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(163..182)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1469..1474,1475..1490,1491..1497)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1505..1521
                     /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     1510..1532
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1606..1623
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1639..2292
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2250..2272
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2299..2322
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2335..2352
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2581..2628
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4221..4676)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4358..4379)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4570..4589
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4718..4740)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4826..4966
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4881..4900
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4981..4998)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5152..5739)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5910..6701,6702..6770))
                     /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     6533..6552
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6771..6875)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg tcttcaagaa
       61 ttcccgacac catcgaatgg tgcaaaacct ttcgcggtat ggcatgatag cgcccggaag
      121 agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg
      181 ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga
      241 aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg
      301 cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc
      361 tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca
      421 gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca
      481 atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg
      541 ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc
      601 agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc
      661 atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct
      721 cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga
      781 tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc
      841 tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg
      901 caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat
      961 acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt
     1021 ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg
     1081 tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca
     1141 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     1201 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat
     1261 taggcacaat tctcatgttt gacagcttat catcgactgc acggtgcacc aatgcttctg
     1321 gcgtcaggca gccatcggaa gctgtggtat ggctgtgcag gtcgtaaatc actgcataat
     1381 tcgtgtcgct caaggcgcac tcccgttctg gataatgttt tttgcgccga catcataacg
     1441 gttctggcaa atattctgaa atgagctgtt gacaattaat catcggctcg tataatgtgt
     1501 ggaattgtga gcggataaca atttcacaca ggaaacagcc agtccgttta ggtgttttca
     1561 cgagcacttc accaacaagg accatagatt atgaaaatcg aagaacacca tcaccatcac
     1621 cattccagcg gtaagcttat gtcccctata ctaggttatt ggaaaattaa gggccttgtg
     1681 caacccactc gacttctttt ggaatatctt gaagaaaaat atgaagagca tttgtatgag
     1741 cgcgatgaag gtgataaatg gcgaaacaaa aagtttgaat tgggtttgga gtttcccaat
     1801 cttccttatt atattgatgg tgatgttaaa ttaacacagt ctatggccat catacgttat
     1861 atagctgaca agcacaacat gttgggtggt tgtccaaaag agcgtgcaga gatttcaatg
     1921 cttgaaggag cggttttgga tattagatac ggtgtttcga gaattgcata tagtaaagac
     1981 tttgaaactc tcaaagttga ttttcttagc aagctacctg aaatgctgaa aatgttcgaa
     2041 gatcgtttat gtcataaaac atatttaaat ggtgatcatg taacccatcc tgacttcatg
     2101 ttgtatgacg ctcttgatgt tgttttatac atggacccaa tgtgcctgga tgcgttccca
     2161 aaattagttt gttttaaaaa acgtattgaa gctatcccac aaattgataa gtacttgaaa
     2221 tccagcaagt atatagcatg gcctttgcag ggctggcaag ccacgtttgg tggtggcgac
     2281 catcctccaa aatcggatct ggaagttctg ttccaggggc ccctgtccag cggtctggtt
     2341 ccgcgtggtt ccggtaccgc ggcccagccg gccatggccc agggcgcgct ggttcgtgtt
     2401 cgtgccctgt atgattacga aggtcaggaa catgatgaac tgtccttcaa agccggtgac
     2461 gaactgacca aaatagaaga tgaagatgaa cagggctggt gtaaaggtcg tctggataac
     2521 ggtcaagttg gtctctatcc agccaactac gttgaagcca ttcaggcggc cgcataataa
     2581 accgatacaa ttaaaggctc cttttggagc cttttttttg gagattttca acgtgaaaaa
     2641 attattattc gcaattcctt tagttgttcc tttctattct cactccgctg aaactgttga
     2701 aagttgttta gcaaaacccc atacagaaaa ttcatttact aacgtctgga aagacgacaa
     2761 aactttagat cgttacgcta actatgaggg ctgtctgtgg aatgctacag gcgttgtagt
     2821 ttgtactggt gacgaaactc agtgtctagc tagagtggcg gtggctctgg ttccggtgat
     2881 tttgattatg aaaagatggc aaacgctaat aagggggcta tgaccgaaaa tgccgatgaa
     2941 aacgcgctac agtctgacgc taaaggcaaa cttgattctg tcgctactga ttacggtgct
     3001 gctatcgatg gtttcattgg tgacgtttcc ggccttgcta atggtaatgg tgctactggt
     3061 gattttgctg gctctaattc ccaaatggct caagtcggtg acggtgataa ttcaccttta
     3121 atgaataatt tccgtcaata tttaccttcc ctccctcaat cggttgaatg tcgccctttt
     3181 gtctttggcg ctggtaaacc atatgaattt tctattgatt gtgacaaaat aaacttattc
     3241 cgtggtgtct ttgcgtttct tttatatgtt gccaccttta tgtatgtatt ttctacgttt
     3301 gctaacatac tgcgtaataa ggagtcttaa tcatgccagt tcttttggct agcgccgccc
     3361 tataccttgt ctgcctcccc gcgttgcgtc gcggtgcatg gagccgggcc acctcgacct
     3421 gaatggaagc cggcggcacc tcgctaacgg attcaccact ccaagaattg gagccaatca
     3481 attcttgcgg agaactgtga atgcgcaaac caacccttgg cagaacatat ccatcgcgtc
     3541 cgccatctcc agcagccgca cgcggcgcat ctcgggcagc gttgggtcct ggccacgggt
     3601 gcgcatgatc gtgctcctgt cgttgaggac ccggctaggc tggcggggtt gccttactgg
     3661 ttagcagaat gaatcaccga tacgcgagcg aacgtgaagc gactgctgct gcaaaacgtc
     3721 tgcgacctga gcaacaacat gaatggtctt cggtttccgt gtttcgtaaa gtctggaaac
     3781 gcggaagtca gcgccctgca ccattatgtt ccggatctgc atcgcaggat gctgctggct
     3841 accctgtgga acacctacat ctgtattaac gaagcgctgg cattgaccct gagtgatttt
     3901 tctctggtcc cgccgcatcc ataccgccag ttgtttaccc tcacaacgtt ccagtaaccg
     3961 ggcatgttca tcatcagtaa cccgtatcgt gagcatcctc tctcgtttca tcggtatcat
     4021 tacccccatg aacagaaatc ccccttacac ggaggcatca gtgaccaaac aggaaaaaac
     4081 cgcccttaac atggcccgct ttatcagaag ccagacatta acgcttctgg agaaactcaa
     4141 cgagctggac gcggatgaac aggcagacat ctgtgaatcg cttcacgacc acgctgatga
     4201 gctttaccgc aggatccgcg aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     4261 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     4321 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     4381 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     4441 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     4501 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     4561 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     4621 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcgcg
     4681 gatcctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     4741 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     4801 cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag
     4861 tgtatactgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg
     4921 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgct cttccgcttc
     4981 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5041 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5101 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5161 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5221 gacaggacta taaagatacc agggtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5281 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5341 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5401 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5461 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5521 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5581 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5641 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5701 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5761 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5821 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5881 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5941 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     6001 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     6061 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     6121 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     6181 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg
     6241 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6301 acatgatccc ccatattgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6361 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6421 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6481 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg ggataatacc
     6541 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6601 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6661 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6721 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6781 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6841 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6901 gacgtctaag aaaccatt
//
