LOCUS       sequence-496798-        5399 bp    DNA     circular SYN 09-JAN-2026
DEFINITION  N-term GST tag E2 variant selected for LUBAC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5399)
  TITLE     Schulman lab plasmids 2025
REFERENCE   2  (bases 1 to 5399)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 9, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: GST-tag LUBAC binding E2 variant D3LU-1
FEATURES             Location/Qualifiers
     source          1..5399
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..654
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     612..634
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             661..681
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(1192..1214)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1359..1377)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1445..1549
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1550..1618,1619..2410)
                     /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(1768..1787)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2581..3169
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3070..3089
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3323..3340
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3413..3490
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3491..4573
                     /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(3510..3529)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4622..4627,4628..4645,4646..4652)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4660..4676
                     /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     4684..4700
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4684..4700
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4696..5223
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4716..4733)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4716..4732)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4725..4747)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(5325..5330,5331..5346,5347..5353)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    5361..5377
                     /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)."
ORIGIN
        1 atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt
       61 ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa
      121 tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat
      181 ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac
      241 atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg
      301 gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt
      361 gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa
      421 acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat
      481 gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa
      541 aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca
      601 tggcctttgc agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat
      661 gaaaacctgt attttcaggg atccatggca ctgaaacgca ttaacaaaga actgagcgat
      721 ctggcacgtg atccgcctgc acagattagc gcaggtccgg ttggtgatga tatgtttcat
      781 tggcaggcaa ccattatggg tccgaatgat agcccgtatc aaggtggtgt tttttttctg
      841 accattcact ttccgaccga ctatccgttt aaaccgccta aagttgcatt taccacgcgt
      901 atttatcatc cgaacattaa tagcaatggc agcatttgtc tggatattct gcgttggtct
      961 tggctgaacg cactgaccat tagcaaagtt ctgctgagca ttgcaagcct gctggatgat
     1021 ccgaatccgg atgatcctct ggttccggaa attgcacgta tctataaaac cgatcgcgat
     1081 aaatacaatc gcattagccg tgaatggacc cagaaatatg caatgtaatg agcggccgca
     1141 tcgtgactga ctgacgatct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     1201 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     1261 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
     1321 tagcgatagc ggagtgtata attcttgaag acgaaagggc ctcgtgatac gcctattttt
     1381 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     1441 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1501 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     1561 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     1621 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     1681 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     1741 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc
     1801 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     1861 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     1921 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     1981 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     2041 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat
     2101 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     2161 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     2221 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     2281 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     2341 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     2401 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     2461 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2521 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2581 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2641 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2701 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     2761 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2821 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2881 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     2941 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3001 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3061 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3121 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3181 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3241 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3301 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     3361 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt ccgacaccat
     3421 cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
     3481 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
     3541 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
     3601 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
     3661 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
     3721 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
     3781 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
     3841 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
     3901 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
     3961 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
     4021 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
     4081 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
     4141 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
     4201 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
     4261 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
     4321 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gtctgctggg
     4381 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     4441 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     4501 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     4561 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcaccccagg
     4621 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     4681 acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt acaacgtcgt
     4741 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     4801 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     4861 aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga aagctggctg
     4921 gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca gatgcacggt
     4981 tacgatgcgc ccatctacac caacgtgacc tatcccatta cggtcaatcc gccgtttgtt
     5041 cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga aagctggcta
     5101 caggaaggcc agacgcgaat tatttttgat ggcgttggaa ttagcttatc gactgcacgg
     5161 tgcaccaatg cttctggcgt caggcagcca tcggaagctg tggtatggct gtgcaggtcg
     5221 taaatcactg cataattcgt gtcgctcaag gcgcactccc gttctggata atgttttttg
     5281 cgccgacatc ataacggttc tggcaaatat tctgaaatga gctgttgaca attaatcatc
     5341 ggctcgtata atgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagtattc
//
