LOCUS       sequence-191015-        6891 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            MYO1F-1/1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6891)
  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 6891)
  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_MYO1F-1/1
FEATURES             Location/Qualifiers
     source          1..6891
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     37..55
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        78..155
                     /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             156..1238
                     /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(175..194)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1481..1486,1487..1502,1503..1509)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1517..1533
                     /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     1522..1544
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1618..1635
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1651..2304
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2262..2284
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2311..2334
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2347..2364
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2566..2613
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4206..4661)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4343..4364)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4555..4574
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4703..4725)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4811..4951
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4866..4885
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4966..4983)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5137..5724)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5895..6686,6687..6755))
                     /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     6518..6537
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6756..6860)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
       61 cgtcttcaag aattcccgac accatcgaat ggtgcaaaac ctttcgcggt atggcatgat
      121 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
      181 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
      241 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
      301 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
      361 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
      421 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
      481 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
      541 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
      601 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
      661 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
      721 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc
      781 aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa
      841 ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga
      901 tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct
      961 cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca
     1021 tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc
     1081 agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca
     1141 ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     1201 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt
     1261 tagctcactc attaggcaca attctcatgt ttgacagctt atcatcgact gcacggtgca
     1321 ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt atggctgtgc aggtcgtaaa
     1381 tcactgcata attcgtgtcg ctcaaggcgc actcccgttc tggataatgt tttttgcgcc
     1441 gacatcataa cggttctggc aaatattctg aaatgagctg ttgacaatta atcatcggct
     1501 cgtataatgt gtggaattgt gagcggataa caatttcaca caggaaacag ccagtccgtt
     1561 taggtgtttt cacgagcact tcaccaacaa ggaccataga ttatgaaaat cgaagaacac
     1621 catcaccatc accattccag cggtaagctt atgtccccta tactaggtta ttggaaaatt
     1681 aagggccttg tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa atatgaagag
     1741 catttgtatg agcgcgatga aggtgataaa tggcgaaaca aaaagtttga attgggtttg
     1801 gagtttccca atcttcctta ttatattgat ggtgatgtta aattaacaca gtctatggcc
     1861 atcatacgtt atatagctga caagcacaac atgttgggtg gttgtccaaa agagcgtgca
     1921 gagatttcaa tgcttgaagg agcggttttg gatattagat acggtgtttc gagaattgca
     1981 tatagtaaag actttgaaac tctcaaagtt gattttctta gcaagctacc tgaaatgctg
     2041 aaaatgttcg aagatcgttt atgtcataaa acatatttaa atggtgatca tgtaacccat
     2101 cctgacttca tgttgtatga cgctcttgat gttgttttat acatggaccc aatgtgcctg
     2161 gatgcgttcc caaaattagt ttgttttaaa aaacgtattg aagctatccc acaaattgat
     2221 aagtacttga aatccagcaa gtatatagca tggcctttgc agggctggca agccacgttt
     2281 ggtggtggcg accatcctcc aaaatcggat ctggaagttc tgttccaggg gcccctgtcc
     2341 agcggtctgg ttccgcgtgg ttccggtacc gcggcccagc cggccccgcg ttgtcgtgct
     2401 ctgtatcagt atgttggtca agatgtcgat gaactgtcct ttaatgtgaa tgaagttatt
     2461 gaaatcctga tggaagatcc gagcggttgg tggaaaggtc gtctgcatgg ccaggaaggt
     2521 ctgtttccgg gcaactatgt ggaaaaaatt gcggccgcat aataaaccga tacaattaaa
     2581 ggctcctttt ggagcctttt ttttggagat tttcaacgtg aaaaaattat tattcgcaat
     2641 tcctttagtt gttcctttct attctcactc cgctgaaact gttgaaagtt gtttagcaaa
     2701 accccataca gaaaattcat ttactaacgt ctggaaagac gacaaaactt tagatcgtta
     2761 cgctaactat gagggctgtc tgtggaatgc tacaggcgtt gtagtttgta ctggtgacga
     2821 aactcagtgt ctagctagag tggcggtggc tctggttccg gtgattttga ttatgaaaag
     2881 atggcaaacg ctaataaggg ggctatgacc gaaaatgccg atgaaaacgc gctacagtct
     2941 gacgctaaag gcaaacttga ttctgtcgct actgattacg gtgctgctat cgatggtttc
     3001 attggtgacg tttccggcct tgctaatggt aatggtgcta ctggtgattt tgctggctct
     3061 aattcccaaa tggctcaagt cggtgacggt gataattcac ctttaatgaa taatttccgt
     3121 caatatttac cttccctccc tcaatcggtt gaatgtcgcc cttttgtctt tggcgctggt
     3181 aaaccatatg aattttctat tgattgtgac aaaataaact tattccgtgg tgtctttgcg
     3241 tttcttttat atgttgccac ctttatgtat gtattttcta cgtttgctaa catactgcgt
     3301 aataaggagt cttaatcatg ccagttcttt tggctagcgc cgccctatac cttgtctgcc
     3361 tccccgcgtt gcgtcgcggt gcatggagcc gggccacctc gacctgaatg gaagccggcg
     3421 gcacctcgct aacggattca ccactccaag aattggagcc aatcaattct tgcggagaac
     3481 tgtgaatgcg caaaccaacc cttggcagaa catatccatc gcgtccgcca tctccagcag
     3541 ccgcacgcgg cgcatctcgg gcagcgttgg gtcctggcca cgggtgcgca tgatcgtgct
     3601 cctgtcgttg aggacccggc taggctggcg gggttgcctt actggttagc agaatgaatc
     3661 accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga cctgagcaac
     3721 aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc
     3781 ctgcaccatt atgttccgga tctgcatcgc aggatgctgc tggctaccct gtggaacacc
     3841 tacatctgta ttaacgaagc gctggcattg accctgagtg atttttctct ggtcccgccg
     3901 catccatacc gccagttgtt taccctcaca acgttccagt aaccgggcat gttcatcatc
     3961 agtaacccgt atcgtgagca tcctctctcg tttcatcggt atcattaccc ccatgaacag
     4021 aaatccccct tacacggagg catcagtgac caaacaggaa aaaaccgccc ttaacatggc
     4081 ccgctttatc agaagccaga cattaacgct tctggagaaa ctcaacgagc tggacgcgga
     4141 tgaacaggca gacatctgtg aatcgcttca cgaccacgct gatgagcttt accgcaggat
     4201 ccgcgaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     4261 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     4321 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     4381 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     4441 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     4501 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     4561 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     4621 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcgcggatcc tgcctcgcgc
     4681 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     4741 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     4801 ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa
     4861 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     4921 cagatgcgta aggagaaaat accgcatcag gcgctcttcc gcttcctcgc tcactgactc
     4981 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5041 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5101 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5161 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5221 ataccagggt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5281 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5341 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5401 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     5461 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5521 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     5581 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5641 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5701 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     5761 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5821 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5881 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5941 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6001 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     6061 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     6121 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6181 aatagtttgc gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg ctcgtcgttt
     6241 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6301 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     6361 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     6421 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     6481 cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga
     6541 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     6601 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6661 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6721 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6781 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6841 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
