LOCUS       sequence-275517-        4997 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of HA-tagged human UFM1-cCS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4997)
  AUTHORS   Liang JR, Lingeman E, Luong T, Ahmed S, Muhar M, Nguyen T, Olzmann 
            JA, Corn JE
  TITLE     A Genome-wide ER-phagy Screen Highlights Key Roles of Mitochondrial 
            Metabolism and ER-Resident UFMylation.
  JOURNAL   Cell. 2020 Mar 19;180(6):1160-1177.e20. doi: 
            10.1016/j.cell.2020.02.017. Epub 2020 Mar 10.
  PUBMED    32160526
REFERENCE   2  (bases 1 to 4997)
  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"; doi: 
            "10.1016/j.cell.2020.02.017"; journalName: "Cell"; date: 
            "2020-03-19- 19"; volume: "180"; issue: "6"; pages: "1160-1177.e20"
COMMENT     Sequence Label: pRK5-HA-UFM1-dCS
FEATURES             Location/Qualifiers
     source          1..4997
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      373..961
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     862..881
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1115..1132
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1249..1270
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1285..1290,1291..1308,1309..1315)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1323..1339
                     /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     1328..1350
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1347..1363
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1347..1363
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        1392..1771
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1772..1975
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1926..1946
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1972..1996
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2206..2224
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2206..2223
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2237..2256
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2324..2350
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2675..2684
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2688..2822
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2725..2744)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2779..2798
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2891..3248
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2891..3082
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2914..2934)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3099..3234
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3268..3285)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3268..3284)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3277..3299)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3497..3952
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3584..3603)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3794..3815
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3969..3988)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4088..4110
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4148..4166)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4234..4338
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4339..4407,4408..4997,1..202)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4557..4576)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
       61 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
      121 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
      181 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
      241 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
      301 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
      361 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
      421 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
      481 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
      541 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
      601 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
      661 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
      721 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
      781 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
      841 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
      901 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
      961 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     1021 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     1081 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     1141 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     1201 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
     1261 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     1321 aattgtgagc ggataacaat ttcacacagg aaacagctat gacatgatta cgaattaatt
     1381 cgagctcgcc cgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     1441 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     1501 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     1561 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     1621 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     1681 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     1741 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     1801 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     1861 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     1921 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctcgttt
     1981 agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc atagaagaca
     2041 ccgggaccga tccagcctcc gcggccggga acggtgcatt ggaacgcgga ttccccgtgc
     2101 caagagtgac gtaagtaccg cctatagagt ctataggccc acccccttgg cttcgttaga
     2161 acgcggctac aattaataca taaccttatg tatcatacac atacgattta ggtgacacta
     2221 tagaataaca tccactttgc ctttctctcc acaggtgtcc actcccaggt ccaactgcac
     2281 ctcggttcta tcgattgaat tccccaccga gctcggatcc atgtacccat acgatgttcc
     2341 agattacgct ggtggatctg gaggttctgg tggaatgtcg aaggtttcct ttaagatcac
     2401 gctgacgtcg gacccacggc tgccgtacaa agtactcagt gttcctgaaa gtacaccttt
     2461 cacagcagtc ttaaagtttg cagcagaaga atttaaagtt cctgctgcaa caagtgcaat
     2521 tattaccaat gatggaatag gaataaatcc tgcacagact gctggaaatg tttttctaaa
     2581 acatggttca gaactgcgga ttattcctag agatcgtgtt ggataatcta gacccagctt
     2641 tggggatcct ctagagtcga cctgcagaag cttggccgcc atggcccaac ttgtttattg
     2701 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     2761 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     2821 tcgggaatta attcggcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt
     2881 taggtacctt ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     2941 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3001 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3061 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3121 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3181 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3241 tttgcaaaaa gctgttaaca gcttggcact ggccgtcgtt ttacaacgtc gtgactggga
     3301 aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg
     3361 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga
     3421 atggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     3481 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     3541 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     3601 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     3661 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
     3721 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     3781 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc
     3841 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     3901 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
     3961 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     4021 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     4081 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     4141 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     4201 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     4261 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     4321 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     4381 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     4441 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     4501 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     4561 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     4621 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     4681 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     4741 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     4801 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     4861 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     4921 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     4981 gcaggaccac ttctgcg
//
