LOCUS       sequence-403938-        5308 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-tagged UFBP1 M2 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5308)
  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 5308)
  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 M2
FEATURES             Location/Qualifiers
     source          1..5308
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(208..213,214..229,230..236)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    244..260
                     /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             283..936
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     894..916
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             943..966
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1383..1405)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1550..1568)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1636..1740
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1741..1809,1810..2601)
                     /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(1959..1978)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2772..3360
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3261..3280
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3514..3531
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3604..3681
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3682..4764
                     /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(3701..3720)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4813..4818,4819..4836,4837..4843)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4851..4867
                     /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     4875..4891
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4875..4891
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4887..5308,1..106)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4907..4924)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4907..4923)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4916..4938)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 aattattttt gatggcgttg gaattagctt atcgactgca cggtgcacca atgcttctgg
       61 cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg tcgtaaatca ctgcataatt
      121 cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt ttgcgccgac atcataacgg
      181 ttctggcaaa tattctgaaa tgagctgttg acaattaatc atcggctcgt ataatgtgtg
      241 gaattgtgag cggataacaa tttcacacag gaaacagtat tcatgtcccc tatactaggt
      301 tattggaaaa ttaagggcct tgtgcaaccc actcgacttc ttttggaata tcttgaagaa
      361 aaatatgaag agcatttgta tgagcgcgat gaaggtgata aatggcgaaa caaaaagttt
      421 gaattgggtt tggagtttcc caatcttcct tattatattg atggtgatgt taaattaaca
      481 cagtctatgg ccatcatacg ttatatagct gacaagcaca acatgttggg tggttgtcca
      541 aaagagcgtg cagagatttc aatgcttgaa ggagcggttt tggatattag atacggtgtt
      601 tcgagaattg catatagtaa agactttgaa actctcaaag ttgattttct tagcaagcta
      661 cctgaaatgc tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt aaatggtgat
      721 catgtaaccc atcctgactt catgttgtat gacgctcttg atgttgtttt atacatggac
      781 ccaatgtgcc tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat tgaagctatc
      841 ccacaaattg ataagtactt gaaatccagc aagtatatag catggccttt gcagggctgg
      901 caagccacgt ttggtggtgg cgaccatcct ccaaaatcgg atctggaagt tctgttccag
      961 gggcccctgg gatccgcatc agccggccaa gagccactgc acaatgagga gctggcagga
     1021 gcaggccggg tggcccagcc tgggcccctg gagcctgagg agccgagagc tggaggcagg
     1081 cctcggcgcc ggagggacct gggcagccgc ctacaggccc agcgtcgagc ccagcgggtg
     1141 gcctgggcag aagcagatga gaacgaggag gaagctgtca tcctagccca ggaggaggaa
     1201 ggtgtcgaga agccagcgga aactcacctg tcggggtgag aattcatgtc gaaggtttcc
     1261 tttaagatca cgctgacgtc ggacccacgg ctgccgtaca aagtactccc gggtcgactc
     1321 gagcggccgc atcgtgactg actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa
     1381 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     1441 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     1501 acccagtcac gtagcgatag cggagtgtat aattcttgaa gacgaaaggg cctcgtgata
     1561 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     1621 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     1681 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     1741 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     1801 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     1861 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     1921 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     1981 cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     2041 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     2101 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     2161 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     2221 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2281 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     2341 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     2401 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     2461 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     2521 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     2581 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     2641 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     2701 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     2761 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     2821 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     2881 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     2941 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3001 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3061 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3121 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3181 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3241 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3301 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3361 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3421 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     3481 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     3541 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat
     3601 tccgacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag
     3661 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
     3721 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
     3781 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
     3841 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
     3901 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
     3961 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
     4021 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
     4081 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
     4141 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
     4201 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
     4261 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
     4321 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     4381 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     4441 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     4501 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     4561 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     4621 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccacgct agcgcccaat
     4681 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     4741 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta
     4801 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     4861 ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt
     4921 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     4981 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     5041 tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg
     5101 aaagctggct ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc
     5161 agatgcacgg ttacgatgcg cccatctaca ccaacgtgac ctatcccatt acggtcaatc
     5221 cgccgtttgt tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg
     5281 aaagctggct acaggaaggc cagacgcg
//
