LOCUS       sequence-191835-        6942 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of human SH3 domain construct 
            CACNB3-1/1-SV1/2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6942)
  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 6942)
  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_CACNB3-1/1-SV1/2
FEATURES             Location/Qualifiers
     source          1..6942
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     8..26
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        49..126
                     /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             127..1209
                     /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(146..165)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1452..1457,1458..1473,1474..1480)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1488..1504
                     /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     1493..1515
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1589..1606
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1622..2275
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2233..2255
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2282..2305
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2318..2335
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      2588..2635
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(4228..4683)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4365..4386)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4577..4596
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4725..4747)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4833..4973
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4888..4907
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4988..5005)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5159..5746)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5917..6708,6709..6777))
                     /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     6540..6559
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6778..6882)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctataaaaat aggcgtatca cgaggccctt tcgtcttcaa gaattcccga caccatcgaa
       61 tggtgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
      121 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
      181 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
      241 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
      301 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
      361 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
      421 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
      481 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
      541 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
      601 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
      661 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
      721 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
      781 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
      841 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
      901 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac
      961 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
     1021 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
     1081 ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
     1141 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     1201 gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac aattctcatg
     1261 tttgacagct tatcatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg
     1321 gaagctgtgg tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg
     1381 cactcccgtt ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct
     1441 gaaatgagct gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata
     1501 acaatttcac acaggaaaca gccagtccgt ttaggtgttt tcacgagcac ttcaccaaca
     1561 aggaccatag attatgaaaa tcgaagaaca ccatcaccat caccattcca gcggtaagct
     1621 tatgtcccct atactaggtt attggaaaat taagggcctt gtgcaaccca ctcgacttct
     1681 tttggaatat cttgaagaaa aatatgaaga gcatttgtat gagcgcgatg aaggtgataa
     1741 atggcgaaac aaaaagtttg aattgggttt ggagtttccc aatcttcctt attatattga
     1801 tggtgatgtt aaattaacac agtctatggc catcatacgt tatatagctg acaagcacaa
     1861 catgttgggt ggttgtccaa aagagcgtgc agagatttca atgcttgaag gagcggtttt
     1921 ggatattaga tacggtgttt cgagaattgc atatagtaaa gactttgaaa ctctcaaagt
     1981 tgattttctt agcaagctac ctgaaatgct gaaaatgttc gaagatcgtt tatgtcataa
     2041 aacatattta aatggtgatc atgtaaccca tcctgacttc atgttgtatg acgctcttga
     2101 tgttgtttta tacatggacc caatgtgcct ggatgcgttc ccaaaattag tttgttttaa
     2161 aaaacgtatt gaagctatcc cacaaattga taagtacttg aaatccagca agtatatagc
     2221 atggcctttg cagggctggc aagccacgtt tggtggtggc gaccatcctc caaaatcgga
     2281 tctggaagtt ctgttccagg ggcccctgtc cagcggtctg gttccgcgtg gttccggtac
     2341 cgcggcccag ccggccatgg cccagggcgc gctgtttgcc gttcgtacca acgtttccta
     2401 ttgcggtgtc ctggatgaag aatgtccagt tcagggttcc ggtgttaact tcgaagccaa
     2461 agattttctg catatcaaag aaaaatactc caacgattgg tggattggtc gtctggttaa
     2521 agaaggtggt gatattgcct tcattccatc cccacaacgt ctggaagaat ctgcggccgc
     2581 ataataaacc gatacaatta aaggctcctt ttggagcctt ttttttggag attttcaacg
     2641 tgaaaaaatt attattcgca attcctttag ttgttccttt ctattctcac tccgctgaaa
     2701 ctgttgaaag ttgtttagca aaaccccata cagaaaattc atttactaac gtctggaaag
     2761 acgacaaaac tttagatcgt tacgctaact atgagggctg tctgtggaat gctacaggcg
     2821 ttgtagtttg tactggtgac gaaactcagt gtctagctag agtggcggtg gctctggttc
     2881 cggtgatttt gattatgaaa agatggcaaa cgctaataag ggggctatga ccgaaaatgc
     2941 cgatgaaaac gcgctacagt ctgacgctaa aggcaaactt gattctgtcg ctactgatta
     3001 cggtgctgct atcgatggtt tcattggtga cgtttccggc cttgctaatg gtaatggtgc
     3061 tactggtgat tttgctggct ctaattccca aatggctcaa gtcggtgacg gtgataattc
     3121 acctttaatg aataatttcc gtcaatattt accttccctc cctcaatcgg ttgaatgtcg
     3181 cccttttgtc tttggcgctg gtaaaccata tgaattttct attgattgtg acaaaataaa
     3241 cttattccgt ggtgtctttg cgtttctttt atatgttgcc acctttatgt atgtattttc
     3301 tacgtttgct aacatactgc gtaataagga gtcttaatca tgccagttct tttggctagc
     3361 gccgccctat accttgtctg cctccccgcg ttgcgtcgcg gtgcatggag ccgggccacc
     3421 tcgacctgaa tggaagccgg cggcacctcg ctaacggatt caccactcca agaattggag
     3481 ccaatcaatt cttgcggaga actgtgaatg cgcaaaccaa cccttggcag aacatatcca
     3541 tcgcgtccgc catctccagc agccgcacgc ggcgcatctc gggcagcgtt gggtcctggc
     3601 cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc
     3661 ttactggtta gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca
     3721 aaacgtctgc gacctgagca acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc
     3781 tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct
     3841 gctggctacc ctgtggaaca cctacatctg tattaacgaa gcgctggcat tgaccctgag
     3901 tgatttttct ctggtcccgc cgcatccata ccgccagttg tttaccctca caacgttcca
     3961 gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag catcctctct cgtttcatcg
     4021 gtatcattac ccccatgaac agaaatcccc cttacacgga ggcatcagtg accaaacagg
     4081 aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca gacattaacg cttctggaga
     4141 aactcaacga gctggacgcg gatgaacagg cagacatctg tgaatcgctt cacgaccacg
     4201 ctgatgagct ttaccgcagg atccgcgaaa ttgtaagcgt taatattttg ttaaaattcg
     4261 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     4321 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     4381 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     4441 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     4501 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     4561 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     4621 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     4681 cgtcgcggat cctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     4741 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag
     4801 ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat
     4861 agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     4921 atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt
     4981 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     5041 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     5101 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     5161 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     5221 gaaacccgac aggactataa agataccagg gtttccccct ggaagctccc tcgtgcgctc
     5281 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5341 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5401 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5461 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5521 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5581 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5641 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     5701 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     5761 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5821 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5881 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5941 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6001 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6061 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6121 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6181 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctgcaggcat
     6241 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6301 gcgagttaca tgatccccca tattgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6361 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6421 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6481 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caacacggga
     6541 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6601 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6661 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     6721 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6781 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6841 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6901 gccacctgac gtctaagaaa ccattattat catgacatta ac
//
