LOCUS       sequence-256097-        5035 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone for AU1 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-1-3xAU1
FEATURES             Location/Qualifiers
     source          1..5035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(512..517,518..535,536..542)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    550..566
                     /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     574..590
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     574..590
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             586..1113
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(606..623)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(606..622)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(615..637)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(1215..1220,1221..1236,1237..1243)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1251..1267
                     /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             1290..1943
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1901..1923
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1950..1967
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2117..2139)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2284..2302)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2370..2474
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2475..2543,2544..3335)
                     /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(2693..2712)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3506..4094
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3995..4014
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4248..4265
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4338..4415
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(4416..5035,1..463)
                     /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(4435..4454)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca
       61 acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga
      121 tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga
      181 tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac
      241 caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact
      301 ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa
      361 aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
      421 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
      481 taagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
      541 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
      601 gattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
      661 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
      721 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcgctttgc ctggtttccg
      781 gcaccagaag cggtgccgga aagctggctg gagtgcgatc ttcctgaggc cgatactgtc
      841 gtcgtcccct caaactggca gatgcacggt tacgatgcgc ccatctacac caacgtgacc
      901 tatcccatta cggtcaatcc gccgtttgtt cccacggaga atccgacggg ttgttactcg
      961 ctcacattta atgttgatga aagctggcta caggaaggcc agacgcgaat tatttttgat
     1021 ggcgttggaa ttagcttatc gactgcacgg tgcaccaatg cttctggcgt caggcagcca
     1081 tcggaagctg tggtatggct gtgcaggtcg taaatcactg cataattcgt gtcgctcaag
     1141 gcgcactccc gttctggata atgttttttg cgccgacatc ataacggttc tggcaaatat
     1201 tctgaaatga gctgttgaca attaatcatc ggctcgtata atgtgtggaa ttgtgagcgg
     1261 ataacaattt cacacaggaa acagtattca tgtcccctat actaggttat tggaaaatta
     1321 agggccttgt gcaacccact cgacttcttt tggaatatct tgaagaaaaa tatgaagagc
     1381 atttgtatga gcgcgatgaa ggtgataaat ggcgaaacaa aaagtttgaa ttgggtttgg
     1441 agtttcccaa tcttccttat tatattgatg gtgatgttaa attaacacag tctatggcca
     1501 tcatacgtta tatagctgac aagcacaaca tgttgggtgg ttgtccaaaa gagcgtgcag
     1561 agatttcaat gcttgaagga gcggttttgg atattagata cggtgtttcg agaattgcat
     1621 atagtaaaga ctttgaaact ctcaaagttg attttcttag caagctacct gaaatgctga
     1681 aaatgttcga agatcgttta tgtcataaaa catatttaaa tggtgatcat gtaacccatc
     1741 ctgacttcat gttgtatgac gctcttgatg ttgttttata catggaccca atgtgcctgg
     1801 atgcgttccc aaaattagtt tgttttaaaa aacgtattga agctatccca caaattgata
     1861 agtacttgaa atccagcaag tatatagcat ggcctttgca gggctggcaa gccacgtttg
     1921 gtggtggcga ccatcctcca aaatcggatc tggttccgcg tggatctatg gacacctacc
     1981 gctacatcga cacctaccgc tacatcgaca cctaccgcta catcaagctt ggatccgaat
     2041 tcccgggtcg actcgagcgg ccgcatcgtg actgactgac gatctgcctc gcgcgtttcg
     2101 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     2161 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     2221 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtataattct tgaagacgaa
     2281 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     2341 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     2401 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     2461 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     2521 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     2581 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     2641 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     2701 gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt
     2761 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     2821 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     2881 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     2941 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     3001 gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac
     3061 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     3121 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     3181 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     3241 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     3301 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     3361 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     3421 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     3481 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     3541 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     3601 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     3661 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     3721 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     3781 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     3841 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     3901 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     3961 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     4021 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     4081 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     4141 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     4201 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     4261 gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     4321 cacaccgcat aaattccgac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat
     4381 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
     4441 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
     4501 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
     4561 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
     4621 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
     4681 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
     4741 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
     4801 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
     4861 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
     4921 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
     4981 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcg
//
