LOCUS       sequence-403798-        5581 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-tagged UFBP1 M3 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5581)
  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 5581)
  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 M3
FEATURES             Location/Qualifiers
     source          1..5581
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(204..209,210..225,226..232)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    240..256
                     /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             279..932
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     890..912
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             939..962
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1652..1674)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1819..1837)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1905..2009
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2010..2078,2079..2870)
                     /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(2228..2247)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3041..3629
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3530..3549
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3783..3800
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3873..3950
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3951..5033
                     /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(3970..3989)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5082..5087,5088..5105,5106..5112)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5120..5136
                     /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     5144..5160
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5144..5160
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5156..5581,1..102)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5176..5193)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5176..5192)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5185..5207)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 atttttgatg gcgttggaat tagcttatcg actgcacggt gcaccaatgc ttctggcgtc
       61 aggcagccat cggaagctgt ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg
      121 tcgctcaagg cgcactcccg ttctggataa tgttttttgc gccgacatca taacggttct
      181 ggcaaatatt ctgaaatgag ctgttgacaa ttaatcatcg gctcgtataa tgtgtggaat
      241 tgtgagcgga taacaatttc acacaggaaa cagtattcat gtcccctata ctaggttatt
      301 ggaaaattaa gggccttgtg caacccactc gacttctttt ggaatatctt gaagaaaaat
      361 atgaagagca tttgtatgag cgcgatgaag gtgataaatg gcgaaacaaa aagtttgaat
      421 tgggtttgga gtttcccaat cttccttatt atattgatgg tgatgttaaa ttaacacagt
      481 ctatggccat catacgttat atagctgaca agcacaacat gttgggtggt tgtccaaaag
      541 agcgtgcaga gatttcaatg cttgaaggag cggttttgga tattagatac ggtgtttcga
      601 gaattgcata tagtaaagac tttgaaactc tcaaagttga ttttcttagc aagctacctg
      661 aaatgctgaa aatgttcgaa gatcgtttat gtcataaaac atatttaaat ggtgatcatg
      721 taacccatcc tgacttcatg ttgtatgacg ctcttgatgt tgttttatac atggacccaa
      781 tgtgcctgga tgcgttccca aaattagttt gttttaaaaa acgtattgaa gctatcccac
      841 aaattgataa gtacttgaaa tccagcaagt atatagcatg gcctttgcag ggctggcaag
      901 ccacgtttgg tggtggcgac catcctccaa aatcggatct ggaagttctg ttccaggggc
      961 ccctgggatc caaaattgga gctaagaaac tgcggaagct ggaggagaaa caagcgcgaa
     1021 aggcccagcg tgaggcagag gaggctgaac gtgaggagcg gaaacgactc gagtcccagc
     1081 gcgaagctga gtggaagaag gaggaggagc ggcttcgcct ggaggaggag cagaaggagg
     1141 aggaggagag gaaggcccgc gaggagcagg cccagcggga gcatgaggag tacctgaaac
     1201 tgaaggaggc ctttgtggtg gaggaggaag gcgtaggaga gaccatgact gaggaacagt
     1261 cccagagctt cctgacagag ttcatcaact acatcaagca gtccaaggtt gtgctcttgg
     1321 aagacctggc ttcccaggtg ggcctacgca ctcaggacac cataaatcgc atccaggacc
     1381 tgctggctga ggggactata acaggtgtga ttgacgaccg gggcaagttc atctacataa
     1441 ccccagagga actggccgcc gtggccaact tcatccgaca gcggggccgg gtgtccatcg
     1501 ccgagcttgc ccaagccagc aactccctca tcgcctgggg ccgggagtcc cctgcccaag
     1561 ccccagcctg agaattcccg ggtcgactcg agcggccgca tcgtgactga ctgacgatct
     1621 gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     1681 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     1741 gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata
     1801 attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     1861 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     1921 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     1981 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2041 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     2101 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     2161 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     2221 aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag caactcggtc
     2281 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     2341 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     2401 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     2461 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     2521 taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac
     2581 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     2641 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     2701 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     2761 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     2821 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     2881 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     2941 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     3001 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     3061 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     3121 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     3181 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     3241 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     3301 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3361 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3421 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3481 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3541 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     3601 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     3661 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     3721 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     3781 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     3841 atctgtgcgg tatttcacac cgcataaatt ccgacaccat cgaatggcgc aaaacctttc
     3901 gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag
     3961 taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg
     4021 tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg
     4081 agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga
     4141 ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta
     4201 aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg
     4261 tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca
     4321 ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc
     4381 cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg
     4441 aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc
     4501 tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat
     4561 atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt
     4621 ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg
     4681 ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg
     4741 ttggtgcgga tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc
     4801 cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct
     4861 tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg
     4921 tgaaaagaaa aaccacgcta gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     4981 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
     5041 gcaattaatg taagttagct cactcattag gcaccccagg ctttacactt tatgcttccg
     5101 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac
     5161 catgattacg gattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt
     5221 tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga
     5281 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcgctttgc
     5341 ctggtttccg gcaccagaag cggtgccgga aagctggctg gagtgcgatc ttcctgaggc
     5401 cgatactgtc gtcgtcccct caaactggca gatgcacggt tacgatgcgc ccatctacac
     5461 caacgtgacc tatcccatta cggtcaatcc gccgtttgtt cccacggaga atccgacggg
     5521 ttgttactcg ctcacattta atgttgatga aagctggcta caggaaggcc agacgcgaat
     5581 t
//
