LOCUS       sequence-275057-        6164 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombinant expression of EIF4G2 with dual-affinity tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6164)
  AUTHORS   Dominguez D, Freese P, Alexis MS, Su A, Hochman M, Palden T, Bazile 
            C, Lambert NJ, Van Nostrand EL, Pratt GA, Yeo GW, Graveley BR, Burge
            CB
  TITLE     Sequence, Structure, and Context Preferences of Human RNA Binding 
            Proteins.
  JOURNAL   Mol Cell. 2018 Jun 7;70(5):854-867.e9. doi: 
            10.1016/j.molcel.2018.05.001. Epub 2018 Jun 7.
  PUBMED    29883606
REFERENCE   2  (bases 1 to 6164)
  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.2018.05.001"; journalName: "Mol Cell"; date: 
            "2018-06-7- 7"; volume: "70"; issue: "5"; pages: "854-867.e9"
COMMENT     Sequence Label: EIF4G2_MIF4G_pGEX
FEATURES             Location/Qualifiers
     source          1..6164
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(539..544,545..562,563..569)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    577..593
                     /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     601..617
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     601..617
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             613..1140
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(633..650)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(633..649)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(642..664)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(1242..1247,1248..1263,1264..1270)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1278..1294
                     /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             1317..1970
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1928..1950
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1977..2000
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2010..2123
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     primer_bind     complement(3273..3295)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3440..3458)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3526..3630
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3631..3699,3700..4491)
                     /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(3849..3868)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4662..5250
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5151..5170
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5404..5421
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5494..5571
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5572..6164,1..490)
                     /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(5591..5610)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg
       61 cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg agggcatcgt
      121 tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc gcgccattac
      181 cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg ataccgaaga
      241 cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc tgctggggca
      301 aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg gcaatcagct
      361 gttgcccgtc tcactggtga aaagaaaaac cacgctagcg cccaatacgc aaaccgcctc
      421 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
      481 cgggcagtga gcgcaacgca attaatgtaa gttagctcac tcattaggca ccccaggctt
      541 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
      601 caggaaacag ctatgaccat gattacggat tcactggccg tcgttttaca acgtcgtgac
      661 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
      721 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
      781 ggcgaatggc gctttgcctg gtttccggca ccagaagcgg tgccggaaag ctggctggag
      841 tgcgatcttc ctgaggccga tactgtcgtc gtcccctcaa actggcagat gcacggttac
      901 gatgcgccca tctacaccaa cgtgacctat cccattacgg tcaatccgcc gtttgttccc
      961 acggagaatc cgacgggttg ttactcgctc acatttaatg ttgatgaaag ctggctacag
     1021 gaaggccaga cgcgaattat ttttgatggc gttggaatta gcttatcgac tgcacggtgc
     1081 accaatgctt ctggcgtcag gcagccatcg gaagctgtgg tatggctgtg caggtcgtaa
     1141 atcactgcat aattcgtgtc gctcaaggcg cactcccgtt ctggataatg ttttttgcgc
     1201 cgacatcata acggttctgg caaatattct gaaatgagct gttgacaatt aatcatcggc
     1261 tcgtataatg tgtggaattg tgagcggata acaatttcac acaggaaaca gtattcatgt
     1321 cccctatact aggttattgg aaaattaagg gccttgtgca acccactcga cttcttttgg
     1381 aatatcttga agaaaaatat gaagagcatt tgtatgagcg cgatgaaggt gataaatggc
     1441 gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct tccttattat attgatggtg
     1501 atgttaaatt aacacagtct atggccatca tacgttatat agctgacaag cacaacatgt
     1561 tgggtggttg tccaaaagag cgtgcagaga tttcaatgct tgaaggagcg gttttggata
     1621 ttagatacgg tgtttcgaga attgcatata gtaaagactt tgaaactctc aaagttgatt
     1681 ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga tcgtttatgt cataaaacat
     1741 atttaaatgg tgatcatgta acccatcctg acttcatgtt gtatgacgct cttgatgttg
     1801 ttttatacat ggacccaatg tgcctggatg cgttcccaaa attagtttgt tttaaaaaac
     1861 gtattgaagc tatcccacaa attgataagt acttgaaatc cagcaagtat atagcatggc
     1921 ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca tcctccaaaa tcggatctgg
     1981 aagttctgtt ccaggggccc ctgggattaa tggacgaaaa aaccaccggt tggcgtggtg
     2041 gtcacgttgt tgaaggtctg gctggtgaac tggaacagct gcgtgctcgt ctggaacacc
     2101 acccgcaggg tcagcgtgaa ccgggatcca tggagagtgc gattgcagaa gggggtgctt
     2161 ctcgtttcag tgcttcttcg ggcggaggag gaagtagggg tgcacctcag cactatccca
     2221 agactgctgg caacagcgag ttcctgggga aaaccccagg gcaaaacgct cagaaatgga
     2281 ttcctgcacg aagcactaga cgagatgaca actccgcagc aaacaactcc gcaaacgaaa
     2341 aagaacgaca tgatgcaatc ttcaggaaag taagaggcat actaaataag cttactcctg
     2401 aaaagtttga caagctatgc cttgagctcc tcaatgtggg tgtagagtct aaactcatcc
     2461 ttaaaggggt catactgctg attgtggaca aagccctaga agagccaaag tatagctcac
     2521 tgtatgctca gctatgtctg cgattggcag aagatgcacc aaactttgat ggcccagcag
     2581 cagagggtca accaggacag aagcaaagca ccacattcag acgcctccta atttccaaat
     2641 tacaagatga atttgaaaac cgaactagaa atgttgatgt ctatgataag cgtgaaaatc
     2701 ccctcctccc cgaggaggag gaacagagag ccattgctaa gatcaagatg ttgggaaaca
     2761 tcaaattcat tggagagctt ggcaagcttg atcttattca cgaatctatc cttcataagt
     2821 gcatcaaaac acttttggaa aagaagaaga gagtccaact caaagatatg ggagaggatt
     2881 tggagtgcct ctgtcagata atgaggacag tgggacctag attagaccat gaacgagcca
     2941 agtccttaat ggatcagtac tttgcccgaa tgtgctcctt gatgttaagt aaggaattgc
     3001 cagcaaggat tcgtttcctg ctgcaggata ccgtagagtt gcgagaacac cattgggttc
     3061 ctcgcaaggc ttttcttgac aatggaccaa agacgatcaa tcaaattcgt caagatgcag
     3121 taaaagatct aggggtgttt attcctgctc ctatggctca agggatgaga agtgacttct
     3181 ttctggaggg accgttcatg ccacccaggt gagcggccgc atcgtgactg actgacgatc
     3241 tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     3301 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     3361 ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat
     3421 aattcttgaa gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata
     3481 ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     3541 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     3601 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     3661 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     3721 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     3781 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     3841 aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt
     3901 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     3961 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     4021 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     4081 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     4141 ataccaaacg acgagcgtga caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa
     4201 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     4261 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     4321 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     4381 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     4441 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     4501 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     4561 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     4621 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     4681 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     4741 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     4801 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     4861 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     4921 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     4981 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     5041 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     5101 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     5161 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     5221 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     5281 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     5341 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     5401 cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg
     5461 catctgtgcg gtatttcaca ccgcataaat tccgacacca tcgaatggcg caaaaccttt
     5521 cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa tgtgaaacca
     5581 gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg
     5641 gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg
     5701 gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg
     5761 attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt
     5821 aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc
     5881 gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc
     5941 attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt
     6001 ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat
     6061 gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg
     6121 ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtct
//
