LOCUS       sequence-256098-        5035 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone for GluGlu tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5035)
  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 5035)
  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: pGEX-4T-1-3xGluGlu
FEATURES             Location/Qualifiers
     source          1..5035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(559..564,565..582,583..589)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    597..613
                     /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     621..637
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     621..637
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             633..1160
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(653..670)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(653..669)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(662..684)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(1262..1267,1268..1283,1284..1290)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1298..1314
                     /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             1337..1990
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1948..1970
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1997..2014
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2018..2035
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     CDS             2036..2053
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     CDS             2054..2071
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     primer_bind     complement(2164..2186)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2331..2349)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2417..2521
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2522..2590,2591..3382)
                     /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(2740..2759)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3553..4141
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4042..4061
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4295..4312
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4385..4462
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(4463..5035,1..510)
                     /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(4482..4501)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag
       61 ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa
      121 atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg
      181 ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg
      241 ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag
      301 gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag
      361 gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg
      421 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
      481 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtaa gttagctcac
      541 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
      601 gagcggataa caatttcaca caggaaacag ctatgaccat gattacggat tcactggccg
      661 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
      721 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
      781 aacagttgcg cagcctgaat ggcgaatggc gctttgcctg gtttccggca ccagaagcgg
      841 tgccggaaag ctggctggag tgcgatcttc ctgaggccga tactgtcgtc gtcccctcaa
      901 actggcagat gcacggttac gatgcgccca tctacaccaa cgtgacctat cccattacgg
      961 tcaatccgcc gtttgttccc acggagaatc cgacgggttg ttactcgctc acatttaatg
     1021 ttgatgaaag ctggctacag gaaggccaga cgcgaattat ttttgatggc gttggaatta
     1081 gcttatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg
     1141 tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt
     1201 ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct
     1261 gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata acaatttcac
     1321 acaggaaaca gtattcatgt cccctatact aggttattgg aaaattaagg gccttgtgca
     1381 acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg
     1441 cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct
     1501 tccttattat attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat
     1561 agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct
     1621 tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt
     1681 tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga
     1741 tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt
     1801 gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa
     1861 attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc
     1921 cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca
     1981 tcctccaaaa tcggatctgg ttccgcgtgg atctatggag tatatgccga tggaggagta
     2041 tatgccgatg gaggagtata tgccgatgga gaagcttgga tccgaattcc cgggtcgact
     2101 cgagcggccg catcgtgact gactgacgat ctgcctcgcg cgtttcggtg atgacggtga
     2161 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     2221 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     2281 gacccagtca cgtagcgata gcggagtgta taattcttga agacgaaagg gcctcgtgat
     2341 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     2401 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2461 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     2521 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     2581 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     2641 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     2701 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     2761 ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     2821 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     2881 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     2941 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     3001 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     3061 gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     3121 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     3181 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     3241 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     3301 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     3361 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     3421 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     3481 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     3541 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     3601 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     3661 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     3721 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     3781 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     3841 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     3901 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     3961 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     4021 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     4081 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     4141 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4201 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     4261 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     4321 agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcataaa
     4381 ttccgacacc atcgaatggc gcaaaacctt tcgcggtatg gcatgatagc gcccggaaga
     4441 gagtcaattc agggtggtga atgtgaaacc agtaacgtta tacgatgtcg cagagtatgc
     4501 cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa
     4561 aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc
     4621 acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct
     4681 gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag
     4741 cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa
     4801 tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc
     4861 cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca
     4921 gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg gcgtggagca
     4981 tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttct
//
