LOCUS       sequence-403794-        5281 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-tagged UFBP1 M4 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5281)
  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 5281)
  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 M4
FEATURES             Location/Qualifiers
     source          1..5281
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(202..207,208..223,224..230)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    238..254
                     /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             277..930
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     888..910
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             937..960
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1350..1372)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1517..1535)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1603..1707
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1708..1776,1777..2568)
                     /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(1926..1945)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2739..3327
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3228..3247
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3481..3498
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3571..3648
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3649..4731
                     /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(3668..3687)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4780..4785,4786..4803,4804..4810)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4818..4834
                     /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     4842..4858
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4842..4858
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4854..5281,1..100)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4874..4891)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4874..4890)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4883..4905)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttttgatggc gttggaatta gcttatcgac tgcacggtgc accaatgctt ctggcgtcag
       61 gcagccatcg gaagctgtgg tatggctgtg caggtcgtaa atcactgcat aattcgtgtc
      121 gctcaaggcg cactcccgtt ctggataatg ttttttgcgc cgacatcata acggttctgg
      181 caaatattct gaaatgagct gttgacaatt aatcatcggc tcgtataatg tgtggaattg
      241 tgagcggata acaatttcac acaggaaaca gtattcatgt cccctatact aggttattgg
      301 aaaattaagg gccttgtgca acccactcga cttcttttgg aatatcttga agaaaaatat
      361 gaagagcatt tgtatgagcg cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg
      421 ggtttggagt ttcccaatct tccttattat attgatggtg atgttaaatt aacacagtct
      481 atggccatca tacgttatat agctgacaag cacaacatgt tgggtggttg tccaaaagag
      541 cgtgcagaga tttcaatgct tgaaggagcg gttttggata ttagatacgg tgtttcgaga
      601 attgcatata gtaaagactt tgaaactctc aaagttgatt ttcttagcaa gctacctgaa
      661 atgctgaaaa tgttcgaaga tcgtttatgt cataaaacat atttaaatgg tgatcatgta
      721 acccatcctg acttcatgtt gtatgacgct cttgatgttg ttttatacat ggacccaatg
      781 tgcctggatg cgttcccaaa attagtttgt tttaaaaaac gtattgaagc tatcccacaa
      841 attgataagt acttgaaatc cagcaagtat atagcatggc ctttgcaggg ctggcaagcc
      901 acgtttggtg gtggcgacca tcctccaaaa tcggatctgg aagttctgtt ccaggggccc
      961 ctgggatcca aaattggagc taagaaactg cggaagctgg aggagaaaca agcgcgaaag
     1021 gcccagcgtg aggcagagga ggctgaacgt gaggagcgga aacgactcga gtcccagcgc
     1081 gaagctgagt ggaagaagga ggaggagcgg cttcgcctgg aggaggagca gaaggaggag
     1141 gaggagagga aggcccgcga ggagcaggcc cagcgggagc atgaggagta cctgaaactg
     1201 aaggaggcct ttgtggtgga ggaggaaggc gtaggagaga ccatgactga ggaacagtcc
     1261 cagagctgag aattcccggg tcgactcgag cggccgcatc gtgactgact gacgatctgc
     1321 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     1381 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     1441 gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtataat
     1501 tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     1561 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1621 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1681 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     1741 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1801 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     1861 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     1921 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
     1981 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     2041 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     2101 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     2161 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     2221 ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta
     2281 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     2341 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     2401 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     2461 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     2521 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     2581 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2641 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2701 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2761 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2821 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2881 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2941 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3001 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3061 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3121 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     3181 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     3241 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     3301 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     3361 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     3421 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     3481 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     3541 ctgtgcggta tttcacaccg cataaattcc gacaccatcg aatggcgcaa aacctttcgc
     3601 ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt gaaaccagta
     3661 acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg
     3721 aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag
     3781 ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc gttgctgatt
     3841 ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa
     3901 tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc
     3961 gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt
     4021 aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac taatgttccg
     4081 gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt ctcccatgaa
     4141 gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg
     4201 ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg gcataaatat
     4261 ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag tgccatgtcc
     4321 ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc gatgctggtt
     4381 gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt
     4441 ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg ttatatcccg
     4501 ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg
     4561 ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg
     4621 aaaagaaaaa ccacgctagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     4681 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     4741 aattaatgta agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     4801 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     4861 tgattacgga ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     4921 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     4981 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgctttgcct
     5041 ggtttccggc accagaagcg gtgccggaaa gctggctgga gtgcgatctt cctgaggccg
     5101 atactgtcgt cgtcccctca aactggcaga tgcacggtta cgatgcgccc atctacacca
     5161 acgtgaccta tcccattacg gtcaatccgc cgtttgttcc cacggagaat ccgacgggtt
     5221 gttactcgct cacatttaat gttgatgaaa gctggctaca ggaaggccag acgcgaatta
     5281 t
//
