LOCUS       sequence-449281-        5773 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mamalian cell expression of SNAP-tag2 in endoplasmic reticulum.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5773)
  AUTHORS   Kuhn S, Nasufovic V, Wilhelm J, Kompa J, de Lange EMF, Lin YH, 
            Egoldt C, Fischer J, Lennoi A, Tarnawski M, Reinstein J, Vlijm R, 
            Hiblot J, Johnsson K
  TITLE     SNAP-tag2 for faster and brighter protein labeling.
  JOURNAL   Nat Chem Biol. 2025 Jul 3. doi: 10.1038/s41589-025-01942-z.
  PUBMED    40610720
REFERENCE   2  (bases 1 to 5773)
  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 Chem 
            Biol. 2025 Jul 3. doi: 10.1038/s41589-025-01942-z."
COMMENT     Sequence Label: pcDNA5/FRT/TO_CalR-SNAP-tag2-KDEL
FEATURES             Location/Qualifiers
     source          1..5773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     387..406
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(625..729)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(804..823)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        995..1374
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1375..1578
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1529..1549
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    1580..1598
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1601..1619
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1629..1653
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     complement(2485..2502)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2491..2715
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2848..2867)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2999..3046
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3054..4076
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3054..4076
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4206..4327
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4243..4262)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4297..4316
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4376..4392)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4376..4392)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4389..4411)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4400..4416
                     /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)."
     promoter        complement(join(4424..4430,4431..4448,4449..4454))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4469..4490
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4607..4624)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4778..5366)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4858..4877)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5537..5773,1..555,556..624))
                     /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"
ORIGIN
        1 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
       61 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
      121 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
      181 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
      241 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
      301 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
      361 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
      421 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
      481 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
      541 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
      601 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
      661 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
      721 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
      781 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
      841 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca
      901 acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg
      961 ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa
     1021 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
     1081 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
     1141 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
     1201 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
     1261 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
     1321 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
     1381 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
     1441 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
     1501 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
     1561 gtctatataa gcagagctct ccctatcagt gatagagatc tccctatcag tgatagagat
     1621 cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt
     1681 gacctccata gaagacaccg ggaccgatcc agcctccgga ctctagcgtt taaacttaag
     1741 cttggtaccg agctcggatc cactagtcca gtgtggtgga attctgcaga tatccagcaa
     1801 ccatgctgct atccgtgccg ttgctgctcg gcctcctcgg cctggccgtc gccggaggca
     1861 gcggaggcag cgaattcgga tccgataaag actgcgaaat gaaacgcact acctatgaca
     1921 gcccgctggg taaactgctg ctgagcggtt gcgagcaggg cctgcaccgc atctatttcg
     1981 tgggcaatgg tcagggcgag cagggtcctc cgggcccgga gccgctgatg caggctaccg
     2041 catggctgaa cgcatatttt caccagccag aggcgattga agagttccct gtaccggccc
     2101 tgcatcaccc ggtatttcag caggaatcct ttacccgtca ggtactgtgg aaactgctga
     2161 aagtagtgaa attcggcgag gtgatctcct attcccagct ggcggcgctg gctggcaacc
     2221 cagcggcaac tgcggcggtg aaaaccgctc tgcgtggtaa tccggtgcca attctgatcc
     2281 cgtgccaccg tgtggttcag ggcgacggcg acgtaggtgg ctacgaaggc cctctgtacg
     2341 tgaaagaatg gctgctggcg cacgaaggtc accgtctggg taaaccaggc ctgggcggcg
     2401 ctccaggatt ctcctcaatt tccgctaaag atgaactgta gtaaccgctc gagtctagag
     2461 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     2521 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     2581 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     2641 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     2701 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     2761 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     2821 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     2881 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     2941 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtacctaga
     3001 agttcctatt ccgaagttcc tattctctag aaagtatagg aacttccttg gccaaaaagc
     3061 ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg
     3121 acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc
     3181 gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt
     3241 atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt ggggaattca
     3301 gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc
     3361 ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat gcgatcgctg
     3421 cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat
     3481 acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa
     3541 ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt
     3601 gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg
     3661 tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg
     3721 attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc
     3781 agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg cggctccggg
     3841 cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg
     3901 atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg
     3961 tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag
     4021 tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaatagcacg
     4081 tactacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4141 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4201 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4261 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4321 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     4381 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4441 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4501 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4561 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4621 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4681 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4741 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4801 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4861 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     4921 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     4981 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5041 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5101 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5161 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5221 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5281 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5341 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5401 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5461 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5521 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5581 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5641 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5701 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5761 tatccgcctc cat
//
