LOCUS       sequence-256101-        6419 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes 3xGluGlu tagged human c-Jun for purification by nickel 
            affinity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6419)
  AUTHORS   Smolko CM, Janes KA
  TITLE     An ultrasensitive fiveplex activity assay for cellular kinases.
  JOURNAL   Sci Rep. 2019 Dec 19;9(1):19409. doi: 10.1038/s41598-019-55998-8.
  PUBMED    31857650
REFERENCE   2  (bases 1 to 6419)
  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"; journalName: "Sci Rep."; 
            date: "2019-12-19"; volume: "9(1):19409. doi"; pages: " 
            10.1038/s41598-019-55998-8"
COMMENT     Sequence Label: pET-29b-3xGluGlu-c-jun-His6
FEATURES             Location/Qualifiers
     source          1..6419
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     921..940
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(960..1037)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1243..1262
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1350..1369
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1350..1368
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1369..1393
                     /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             1408..1430
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1441..1485
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             1501..1518
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1522..1539
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     CDS             1540..1557
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     CDS             1558..1575
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     CDS             complement(1606..1617)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             2629..2646
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2699..2717)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2713..2760
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2797..3252
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2884..2903)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3094..3115
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3344..4159)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3344..4159)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4067..4086
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4151..4170
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4281..4869
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4770..4789
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5023..5040
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5055..5194
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5120..5139)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5280..5302
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5296..5487)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6296..6419,1..959))
                     /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"
ORIGIN
        1 ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg
       61 ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat
      121 gagctgtctt cggtatcgtc gtatcccact accgagatgt ccgcaccaac gcgcagcccg
      181 gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca
      241 gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc
      301 cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag
      361 ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc
      421 tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa
      481 ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg
      541 caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca
      601 ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt
      661 tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg
      721 acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac
      781 tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc
      841 gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa
      901 acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca
      961 ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg
     1021 cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa
     1081 gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa
     1141 ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca
     1201 agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata
     1261 ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag
     1321 gatcgagatc gatctcgatc ccgcgaaatt aatacgactc actatagggg aattgtgagc
     1381 ggataacaat tcccctctag aaataatttt gtttaacttt aagaaggaga tatacatatg
     1441 aaagaaaccg ctgctgctaa attcgaacgc cagcacatgg acagcccaga tctgggtacc
     1501 ctggtgccac gcggttccat ggagtatatg ccgatggagg agtatatgcc gatggaggag
     1561 tatatgccga tggagggatc cgaattcatg actgcaaaga tggaaacgac cttctatgac
     1621 gatgccctca acgcctcgtt cctcccgtcc gagagcggac cttatggcta cagtaacccc
     1681 aagatcctga aacagagcat gaccctgaac ctggccgacc cagtggggag cctgaagccg
     1741 cacctccgcg ccaagaactc ggacctcctc acctcgcccg acgtggggct gctcaagctg
     1801 gcgtcgcccg agctggagcg cctgataatc cagtccagca acgggcacat caccaccacg
     1861 ccgaccccca cccagttcct gtgccccaag aacgtgacag atgagcagga gggcttcgcc
     1921 gagggcttcg tgcgcgccct ggccgaactg cacagccaga acacgctgcc cagcgtcacg
     1981 tcggcggcgc agccggtcaa cggggcaggc atggtggctc ccgcggtagc ctcggtggca
     2041 gggggcagcg gcagcggcgg cttcagcgcc agcctgcaca gcgagccgcc ggtctacgca
     2101 aacctcagca acttcaaccc aggcgcgctg agcagcggcg gcggggcgcc ctcctacggc
     2161 gcggccggcc tggcctttcc cgcgcaaccc cagcagcagc agcagccgcc gcaccacctg
     2221 ccccagcaga tgcccgtgca gcacccgcgg ctgcaggccc tgaaggagga gcctcagaca
     2281 gtgcccgaga tgcccggcga gacaccgccc ctgtccccca tcgacatgga gtcccaggag
     2341 cggatcaagg cggagaggaa gcgcatgagg aaccgcatcg ctgcctccaa gtgccgaaaa
     2401 aggaagctgg agagaatcgc ccggctggag gaaaaagtga aaaccttgaa agctcagaac
     2461 tcggagctgg cgtccacggc caacatgctc agggaacagg tggcacagct taaacagaaa
     2521 gtcatgaacc acgttaacag tgggtgccaa ctcatgctaa cgcagcagtt gcaaacattt
     2581 ggatccgaat tcgcgagctc cgtcgacaag cttgcggccg cactcgagca ccaccaccac
     2641 caccactgag atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc
     2701 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     2761 ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa
     2821 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     2881 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     2941 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     3001 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     3061 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     3121 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     3181 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     3241 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     3301 tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa aactcatcga
     3361 gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa
     3421 gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct
     3481 ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt
     3541 caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg
     3601 gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat
     3661 caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa
     3721 atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga
     3781 acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga
     3841 atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa
     3901 aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat
     3961 ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg
     4021 gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt
     4081 tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag caagacgttt
     4141 cccgttgaat atggctcata acaccccttg tattactgtt tatgtaagca gacagtttta
     4201 ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     4261 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     4321 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     4381 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     4441 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     4501 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     4561 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4621 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     4681 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     4741 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     4801 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     4861 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     4921 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4981 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     5041 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     5101 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     5161 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     5221 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     5281 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     5341 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     5401 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     5461 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     5521 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     5581 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     5641 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     5701 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     5761 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     5821 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     5881 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     5941 catgcgcacc cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt
     6001 ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag
     6061 cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag
     6121 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
     6181 gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat
     6241 cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact
     6301 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     6361 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacg
//
