LOCUS       Exported                7651 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expression of human eEF2K in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3-HA-eEF2K
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7651)
  AUTHORS   Kruiswijk F, Yuniati L, Magliozzi R, Low TY, Lim R, Bolder R, 
            Mohammed S, Proud CG, Heck AJ, Pagano M, Guardavaccaro D
  TITLE     Coupled activation and degradation of eEF2K regulates protein 
            synthesis in response to genotoxic stress.
  JOURNAL   Sci Signal. 2012 Jun 5;5(227):ra40. doi: 10.1126/scisignal.2002718.
  PUBMED    22669845
REFERENCE   2  (bases 1 to 7651)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7651
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        618..821
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     866..885
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        866..884
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             929..955
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(3245..3262)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3251..3475
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3521..3949
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3608..3627)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3818..3839
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3958..3978)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3963..4292
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4143..4278
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4205..4224
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4359..5153
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4413..4432)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5023..5042
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5327..5448
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5364..5383)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5418..5437
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5497..5513)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5497..5513)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5510..5532)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5521..5537
                     /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)."
     promoter        complement(5545..5575)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5590..5611
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5728..5745)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5899..6487)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5979..5998)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6658..7518)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7281..7300
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7519..7623)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tctctggcta actagagaac
      841 ccactgctta ctggcttatc gaaattaata cgactcacta tagggagacc caagctggct
      901 agcgtttaaa cttaagctta ccatgggata cccatacgat gttccagatt acgctgtggt
      961 acctaggatc cagtgtggtg gaattcggct tatggcagac gaagacctca tcttccgcct
     1021 ggaaggcgtt gatggcggcc agtccccccg agctggccat gatggtgatt ctgatgggga
     1081 cagcgacgat gaggaaggtt acttcatctg ccccatcacg gatgacccaa gctcgaacca
     1141 gaatgtcaat tccaaggtta ataagtacta cagcaaccta acaaaaagtg agcggtatag
     1201 ctccagcggg tccccggcaa actccttcca cttcaaggaa gcctggaagc acgcaatcca
     1261 gaaggccaag cacatgcccg acccctgggc tgagttccac ctggaagata ttgccaccga
     1321 acgtgctact cgacacaggt acaacgccgt caccggggaa tggctggatg atgaagttct
     1381 gatcaagatg gcatctcagc ccttcggccg aggagcaatg agggagtgct tccggacgaa
     1441 gaagctctcc aacttcttgc atgcccagca gtggaaggga gcctccaact acgtggcgaa
     1501 gcgctacatc gagcccgtag accgggatgt gtactttgag gacgtgcgtc tacagatgga
     1561 ggccaagctc tggggggagg agtataatcg gcacaagccc cccaagcagg tggacatcat
     1621 gcagatgtgc atcatcgagc tgaaggacag accgggcaag cccctcttcc acctggagca
     1681 ctacatcgag ggcaagtaca tcaagtacaa ctccaactct ggctttgtcc gcgatgacaa
     1741 catccgcctg acgccgcagg ccttcagcca cttcactttt gagcgttccg gccatcagct
     1801 gatagtggtg gacatccagg gagttgggga tctctacact gacccacaga tccacacgga
     1861 gacgggcact gactttggag acggcaacct aggtgtccgc gggatggcgc tcttcttcta
     1921 ctctcatgcc tgcaaccgga tttgcgagag catgggcctt gctccctttg acctctcgcc
     1981 ccgggagagg gatgcagtga atcagaacac caagctgctg caatcagcca agaccatctt
     2041 gagaggaaca gaggaaaaat gtgggagccc ccgagtaagg accctctctg ggagccggcc
     2101 acccctgctc cgtccccttt cagagaactc tggagacgag aacatgagcg acgtgacctt
     2161 cgactctctc ccttcttccc catcttcggc cacaccacac agccagaagc tagaccacct
     2221 ccattggcca gtgttcagtg acctcgataa catggcatcc agagaccatg atcatctaga
     2281 caaccaccgg gagtctgaga atagtgggga cagcggatac cccagtgaga agcggggtga
     2341 gctggatgac cctgagcccc gagaacatgg ccactcatac agtaatcgga agtacgagtc
     2401 tgacgaagac agcctgggca gctctggacg ggtatgtgta gagaagtgga atctcctcaa
     2461 ctcctcccgc ctccacctgc cgagggcttc ggccgtggcc ctggaagtgc aaaggcttaa
     2521 tgctctggac ctcgaaaaga aaatcgggaa gtccattttg gggaaggtcc atctggccat
     2581 ggtgcgctac cacgagggtg ggcgcttctg cgagaagggc gaggagtggg accaggagtc
     2641 ggctgtcttc cacctggagc acgcagccaa cctgggcgag ctggaggcca tcgtgggcct
     2701 ggggctcatg tactcgcagt tgcctcatca catcctagcc gatgtctctc tgaaggagac
     2761 agaagagaac aaaaccaaag gatttgatta cttactaaag gccgctgaag ctggcgacag
     2821 gcagtccatg atcctagtgg cgcgagcttt tgactctggc cagaacctca gcccggacag
     2881 gtgccaagac tggctagagg ccctgcactg gtacaacact gccctggaga tgacggactg
     2941 tgatgagggc ggtgagtacg acggaatgca ggacgagccc cggtacatga tgctggccag
     3001 ggaggccgag atgctgttca caggaggcta cgggctggag aaggacccgc agagatcagg
     3061 ggacttgtat acccaggcag cagaggcagc gatggaagcc atgaagggcc gactggccaa
     3121 ccagtactac caaaaggctg aagaggcctg ggcccaaatg gaggaataaa agccgaattc
     3181 tgcagatatc cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga
     3241 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     3301 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     3361 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     3421 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     3481 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     3541 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     3601 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3661 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3721 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3781 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3841 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3901 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     3961 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4021 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     4081 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     4141 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     4201 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     4261 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     4321 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     4381 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     4441 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     4501 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     4561 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     4621 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     4681 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     4741 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     4801 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     4861 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     4921 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     4981 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     5041 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     5101 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     5161 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     5221 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5281 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     5341 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     5401 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     5461 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     5521 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     5581 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     5641 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5701 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     5761 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     5821 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     5881 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     5941 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     6001 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6061 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     6121 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6181 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6241 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6301 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     6361 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6421 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6481 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6541 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     6601 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     6661 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     6721 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     6781 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6841 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     6901 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     6961 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     7021 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     7081 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     7141 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     7201 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     7261 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     7321 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     7381 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     7441 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7501 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7561 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     7621 gcgcacattt ccccgaaaag tgccacctga c
//
