LOCUS       sequence-356605-        7113 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of DmeIF4G_601-660.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7113)
  AUTHORS   Gruner S, Peter D, Weber R, Wohlbold L, Chung MY, Weichenrieder O, 
            Valkov E, Igreja C, Izaurralde E
  TITLE     The Structures of eIF4E-eIF4G Complexes Reveal an Extended Interface
            to Regulate Translation Initiation.
  JOURNAL   Mol Cell. 2016 Nov 3;64(3):467-479. doi: 
            10.1016/j.molcel.2016.09.020. Epub 2016 Oct 20.
  PUBMED    27773676
REFERENCE   2  (bases 1 to 7113)
  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.molcel.2016.09.020"; journalName: "Mol Cell"; date: 
            "2016-11-3- 3"; volume: "64"; issue: "3"; pages: "467-479"
COMMENT     Sequence Label: pnEA-NpM-DmeIF4G_601-660-GB1_AD
FEATURES             Location/Qualifiers
     source          1..7113
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             88..1188
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="METEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     CDS             1204..1227
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1615..1633)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1629..1676
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1851..1856,1857..1873,1874..1879))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1906..1924)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1992..2096
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2097..2165,2166..2957)
                     /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(2315..2334)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3128..3716
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3617..3636
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3870..3887
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3902..4041
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3967..3986)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4127..4149
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4143..4334)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     6689..6708
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6728..6805)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7011..7030
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttccccacta gtaataattt
       61 tgtttaactt taagaaggag atataccatg gaaactgaag aaggtaaact ggtaatctgg
      121 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
      181 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
      241 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
      301 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
      361 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
      421 gaagcgttat cgctgattta taacaaagac ctgctgccga acccgccaaa aacctgggaa
      481 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgccct gatgttcaac
      541 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
      601 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
      661 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
      721 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
      781 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
      841 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
      901 ccgaacaaag agctggcgaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
      961 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     1021 ttggcgaaag atccacgtat tgccgccacg atggaaaacg cccagaaagg tgaaatcatg
     1081 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     1141 gccagcggtc gtcagactgt cgatgaagcc cttaaagacg cgcagactgg taccggcagc
     1201 ggcctggaag ttctgttcca ggggccccat atgtctatca taaactataa tgagggtcaa
     1261 tggtcaccaa acaatccaag cggaaaaaaa caatacgacc gagaacagtt attacagtta
     1321 cgcgaggtta aggcctcacg tatacaacca gaggtaaaaa atgtgtctat tcttcctcaa
     1381 ccaaacttaa tgccatcatt tattcgaaat aacggatccg ctagcggcat gcagtacaaa
     1441 cttatcctga acggtaaaac cctgaaaggt gaaaccacca ccgaagctgt tgacgctgct
     1501 accgcggaaa aagttttcaa acagtacgct aacgacaacg gtgttgacgg tgaatggacc
     1561 tacgacgacg ctaccaaaac ctacaccgta acggaatgag ttggctgctg ccacccgctg
     1621 agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga
     1681 aaggaggaac tatatccgga tatcccgcaa gaggcccggc agtaccggca taaccaagcc
     1741 tatgcctaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat tgttagattt
     1801 catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc attaaagctt
     1861 atcgatgata agctgtcaaa catgagaatt cttgaagacg aaagggcctc gtgatacgcc
     1921 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     1981 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2041 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     2101 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     2161 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     2221 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     2281 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     2341 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     2401 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     2461 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     2521 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     2581 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     2641 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     2701 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     2761 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     2821 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     2881 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     2941 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     3001 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     3061 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     3121 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     3181 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     3241 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     3301 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     3361 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     3421 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     3481 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     3541 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     3601 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3661 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3721 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     3781 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     3841 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     3901 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     3961 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     4021 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     4081 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     4141 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     4201 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     4261 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     4321 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     4381 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     4441 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     4501 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     4561 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     4621 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     4681 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     4741 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     4801 ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc tgctggagat ggcggacgcg
     4861 atggatatgt tctgccaagg gttggtttgc gcattcacag ttctccgcaa gaattgattg
     4921 gctccaattc ttggagtggt gaatccgtta gcgaggtgcc gccggcttcc attcaggtcg
     4981 aggtggcccg gctccatgca ccgcgacgca acgcggggag gcagacaagg tatagggcgg
     5041 cgcctacaat ccatgccaac ccgttccatg tgctcgccga ggcggcataa atcgccgtga
     5101 cgatcagcgg tccaatgatc gaagttaggc tggtaagagc cgcgagcgat ccttgaagct
     5161 gtccctgatg gtcgtcatct acctgcctgg acagcatggc ctgcaacgcg ggcatcccga
     5221 tgccgccgga agcgagaaga atcataatgg ggaaggccat ccagcctcgc gtcgcgaacg
     5281 ccagcaagac gtagcccagc gcgtcggccg ccatgccggc gataatggcc tgcttctcgc
     5341 cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga
     5401 ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa
     5461 tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa
     5521 gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc
     5581 tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt
     5641 tgcgctcact gcccgctttc cagtcgggaa actgtcgtgc cagctgcatt aatgaatcgg
     5701 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca
     5761 gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc
     5821 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
     5881 tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc
     5941 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
     6001 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
     6061 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
     6121 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
     6181 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat
     6241 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
     6301 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
     6361 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
     6421 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
     6481 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
     6541 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
     6601 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
     6661 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
     6721 gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
     6781 aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc
     6841 attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt
     6901 gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg
     6961 ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg
     7021 gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg
     7081 gcgtagagga tcgagatctc gatcccgcga aat
//
