LOCUS       sequence-408411-        5284 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-tagged UFBP1 M5 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5284)
  AUTHORS   Ishimura R, El-Gowily AH, Noshiro D, Komatsu-Hirota S, Ono Y, Shindo
            M, Hatta T, Abe M, Uemura T, Lee-Okada HC, Mohamed TM, Yokomizo T, 
            Ueno T, Sakimura K, Natsume T, Sorimachi H, Inada T, Waguri S, Noda 
            NN, Komatsu M
  TITLE     The UFM1 system regulates ER-phagy through the ufmylation of CYB5R3.
  JOURNAL   Nat Commun. 2022 Dec 21;13(1):7857. doi: 10.1038/s41467-022-35501-0.
  PUBMED    36543799
REFERENCE   2  (bases 1 to 5284)
  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: "Nat 
            Commun."; date: "2022-12-21"; volume: "13(1):7857. doi"; pages: " 
            10.1038/s41467-022-35501-0"
COMMENT     Sequence Label: pGEX6p-1-UFBP1 M5
FEATURES             Location/Qualifiers
     source          1..5284
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1334..1356)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1501..1519)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1587..1691
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1692..1760,1761..2552)
                     /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(1910..1929)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2723..3311
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3212..3231
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3465..3482
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3555..3632
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3633..4715
                     /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(3652..3671)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4764..4769,4770..4787,4788..4794)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4802..4818
                     /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     4826..4842
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4826..4842
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4838..5284,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4858..4875)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4858..4874)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4867..4889)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc ttcctgacag
      961 agttcatcaa ctacatcaag cagtccaagg ttgtgctctt ggaagacctg gcttcccagg
     1021 tgggcctacg cactcaggac accataaatc gcatccagga cctgctggct gaggggacta
     1081 taacaggtgt gattgacgac cggggcaagt tcatctacat aaccccagag gaactggccg
     1141 ccgtggccaa cttcatccga cagcggggcc gggtgtccat cgccgagctt gcccaagcca
     1201 gcaactccct catcgcctgg ggccgggagt cccctgccca agccccagcc tgagaattcc
     1261 cgggtcgact cgagcggccg catcgtgact gactgacgat ctgcctcgcg cgtttcggtg
     1321 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     1381 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     1441 gcgcagccat gacccagtca cgtagcgata gcggagtgta taattcttga agacgaaagg
     1501 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     1561 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     1621 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1681 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1741 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1801 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1861 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     1921 cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     1981 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     2041 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     2101 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     2161 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     2221 acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     2281 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     2341 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     2401 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     2461 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     2521 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     2581 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     2641 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2701 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2761 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2821 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     2881 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2941 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3001 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3061 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3121 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3181 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3241 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3301 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3361 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3421 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3481 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3541 accgcataaa ttccgacacc atcgaatggc gcaaaacctt tcgcggtatg gcatgatagc
     3601 gcccggaaga gagtcaattc agggtggtga atgtgaaacc agtaacgtta tacgatgtcg
     3661 cagagtatgc cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg
     3721 tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca
     3781 accgcgtggc acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca
     3841 gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac
     3901 tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg
     3961 cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg
     4021 accaggatgc cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg
     4081 tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg
     4141 gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa
     4201 gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa
     4261 ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca
     4321 tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg
     4381 cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg
     4441 tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca
     4501 aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg
     4561 gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccacgc
     4621 tagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
     4681 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtaagttag
     4741 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
     4801 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cggattcact
     4861 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     4921 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     4981 ttcccaacag ttgcgcagcc tgaatggcga atggcgcttt gcctggtttc cggcaccaga
     5041 agcggtgccg gaaagctggc tggagtgcga tcttcctgag gccgatactg tcgtcgtccc
     5101 ctcaaactgg cagatgcacg gttacgatgc gcccatctac accaacgtga cctatcccat
     5161 tacggtcaat ccgccgtttg ttcccacgga gaatccgacg ggttgttact cgctcacatt
     5221 taatgttgat gaaagctggc tacaggaagg ccagacgcga attatttttg atggcgttgg
     5281 aatt
//
