LOCUS       sequence-289204-        5599 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial Expression of HsSmg6_1-207-EBM2mut.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5599)
  AUTHORS   Kashima I, Jonas S, Jayachandran U, Buchwald G, Conti E, Lupas AN, 
            Izaurralde E
  TITLE     SMG6 interacts with the exon junction complex via two conserved 
            EJC-binding motifs (EBMs) required for nonsense-mediated mRNA decay.
  JOURNAL   Genes Dev. 2010 Nov 1. 24(21):2440-50.
  PUBMED    20930030
REFERENCE   2  (bases 1 to 5599)
  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"; journalName: "Genes Dev. 
            2010 Nov 1. 24(21):2440-50."
COMMENT     Sequence Label: pGEX-6P-1-HsSmg6_1-207-EBM2mut_K
FEATURES             Location/Qualifiers
     source          1..5599
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1649..1671)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1816..1834)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1902..2006
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2007..2075,2076..2867)
                     /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     complement(2225..2244)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3038..3626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3780..3797
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3870..3947
                     /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             3948..5030
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3967..3986)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5043..5064
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5079..5084,5085..5102,5103..5109)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5117..5133
                     /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     5141..5157
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5141..5157
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5153..5599,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5173..5190)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5173..5189)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5182..5204)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc ccggaattga
      961 tggcggaagg gctggagcgt gtgcggatct ccgcgtcgga gctgcgcggg atcctggcta
     1021 ctctggcccc gcaggccggg agcagagaaa acatgaagga attaaaggag gccaggccgc
     1081 gcaaagataa caggcgtcca gatctggaaa tctataagcc tggcctttct cggctaagga
     1141 acaagcccaa aatcaaggaa ccccctggga gtgaggaatt caaagatgaa attgttaatg
     1201 accgagattg ctctgctgtt gaaaatggta cacagcccgt taaagatgtc tgcaaggaac
     1261 tgaacaacca agagcagaat ggtcctatag acccagaaaa taatcgggga caagaatcct
     1321 ttcctaggac tgctggacaa gaggatcgta gtctaaaaat tatcaaaaga acaaaggaac
     1381 ccgacctgca gatcgaacag cctggacgac gtttgcagac tgttagcaaa gaatccgcca
     1441 gtcgggtgga ggaggaagaa gtcctcaacc aggtagaaca actgagagta gaggaagatg
     1501 agtgtagggg aaatgttgcg aaggaggaag ttgcgaataa accagacagg gccgagatag
     1561 aaaagagccc aggtggtggg tagctcgagc ggccgcatcg tgactgactg acgatctgcc
     1621 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     1681 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     1741 ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtataatt
     1801 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     1861 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     1921 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     1981 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     2041 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     2101 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     2161 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     2221 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     2281 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     2341 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     2401 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     2461 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     2521 caaacgacga gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat
     2581 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     2641 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     2701 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     2761 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     2821 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     2881 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     2941 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3001 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3061 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3121 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3181 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3241 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3301 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3361 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     3421 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     3481 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     3541 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     3601 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     3661 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     3721 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     3781 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     3841 tgtgcggtat ttcacaccgc ataaattccg acaccatcga atggtgcaaa acctttcgcg
     3901 gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa
     3961 cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga
     4021 accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc
     4081 tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg
     4141 gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat
     4201 ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg
     4261 aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta
     4321 actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg
     4381 cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag
     4441 acggtacgcg actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt
     4501 tagcgggccc attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc
     4561 tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg
     4621 gttttcaaca aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg
     4681 ccaacgatca gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg
     4741 gtgcggatat ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc
     4801 cgtcaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc
     4861 tgcaactctc tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga
     4921 aaagaaaaac caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     4981 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     5041 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     5101 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
     5161 gattacggat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     5221 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     5281 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gctttgcctg
     5341 gtttccggca ccagaagcgg tgccggaaag ctggctggag tgcgatcttc ctgaggccga
     5401 tactgtcgtc gtcccctcaa actggcagat gcacggttac gatgcgccca tctacaccaa
     5461 cgtaacctat cccattacgg tcaatccgcc gtttgttccc acggagaatc cgacgggttg
     5521 ttactcgctc acatttaatg ttgatgaaag ctggctacag gaaggccaga cgcgaattat
     5581 ttttgatggc gttggaatt
//
