LOCUS       sequence-205148-        5144 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  T7 promotor drives in vitro transcription of mCherry-tagged wildtype
            mouse TRIM21 mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5144)
  AUTHORS   Clift D, McEwan WA, Labzin LI, Konieczny V, Mogessie B, James LC, 
            Schuh M
  TITLE     A Method for the Acute and Rapid Degradation of Endogenous Proteins.
  JOURNAL   Cell. 2017 Nov 13. pii: S0092-8674(17)31255-2. doi: 
            10.1016/j.cell.2017.10.033.
  PUBMED    29153837
REFERENCE   2  (bases 1 to 5144)
  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: "Cell. 2017 
            Nov 13. pii: S0092-8674(17)31255-2. doi: 
            10.1016/j.cell.2017.10.033."
COMMENT     Sequence Label: pGEMHE-mCherry-mTrim21
FEATURES             Location/Qualifiers
     source          1..5144
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        complement(52..70)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(53..70)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(88..104)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(88..104)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(101..123)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    112..128
                     /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(136..142,143..160,161..166))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..202
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(319..336)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(490..1078)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(570..589)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1249..2040,2041..2109))
                     /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     1872..1891
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2110..2214)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2282..2300
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2338..2360)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2460..2479
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2673..2695
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2687..2704
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2688..2704
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2711..2730
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2711..2729
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     5'UTR           2761..2803
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     regulatory      2837..2846
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2843..3550
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2988..3006)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3256..3275
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3587..4975
                     /codon_start=1
                     /product="mouse RING E3 ubiquitin ligase that can be used 
                     together with antibodies to degrade intracellular proteins 
                     (Clift et al., 2018)"
                     /label=TRIM21
                     /note="binds to the Fc domain of antibodies"
                     /translation="MSLEKMWEEVTCSICLDPMVEPMSIECGHCFCKECIFEVGKNGGS
                     SCPECRQQFLLRNLRPNRHIANMVENLKQIAQNTKKSTQETHCMKHGEKLHLFCEEDGQ
                     ALCWVCAQSGKHRDHTRVPIEEAAKVYQEKIHVALEKLRKGKELAEKMEMDLTMQRTDW
                     KRNIDTQKSRIHAEFALQNSLLAQEEQRQLQRLEKDQREYLRLLGKKEAELAEKNQALQ
                     ELISELERRIRGSELELLQEVRIILERSGSWNLDTLDIDAPDLTSTCPVPGRKKMLRTC
                     WVHITLDRNTANSWLIISKDRRQVRMGDTHQNVSDNKERFSNYPMVLGAQRFSSGKMYW
                     EVDVTQKEAWDLGVCRDSVQRKGQFSLSPENGFWTIWLWQDSYEAGTSPQTTLHIQVPP
                     CQIGIFVDYEAGVVSFYNITDHGSLIYTFSECVFAGPLRPFFNVGFNYSGGNAAPLKLC
                     PLKM"
     3'UTR           5019..5144
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
ORIGIN
        1 ccctgcaggc gcgccgccgg cgatttaaat taattaatct agcttgagta ttctatagtg
       61 tcacctaaat agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
      121 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
      181 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
      241 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
      301 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
      361 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
      421 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
      481 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
      541 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
      601 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
      661 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
      721 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
      781 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
      841 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
      901 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
      961 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     1021 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     1081 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     1141 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     1201 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     1261 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     1321 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     1381 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     1441 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     1501 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     1561 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     1621 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     1681 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     1741 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     1801 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     1861 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     1921 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     1981 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     2041 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     2101 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     2161 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     2221 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     2281 aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct
     2341 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     2401 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc
     2461 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     2521 cgtaaggaga aaataccgca tcaggcgcca ttcgccattc aggctgcgca actgttggga
     2581 agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc
     2641 aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc
     2701 cagtgaattg taatacgact cactataggg cgaattaatt cgagctcggt acccagcttg
     2761 cttgttcttt ttgcagaagc tcagaataaa cgctcaactt tggcagatca attccccggt
     2821 gctagcgcta ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat
     2881 caaggagttc atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga
     2941 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
     3001 gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg
     3061 ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc
     3121 cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac
     3181 ccaggactcc tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa
     3241 cttcccctcc gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga
     3301 gcggatgtac cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa
     3361 ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca
     3421 gctgcccggc gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta
     3481 caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga
     3541 gctgtacaag tccggactca gatctcgagc tcaagcttcg aattcaatgt ctctggaaaa
     3601 gatgtgggag gaggtcacct gttctatctg cctggatccc atggtggagc ctatgagtat
     3661 cgaatgtggc cattgctttt gcaaggaatg catttttgaa gttgggaaga atgggggcag
     3721 ttcatgtccc gagtgccggc aacagtttct gctccgaaac ctcaggccca atagacatat
     3781 agccaacatg gtggaaaacc ttaaacagat agcccagaat accaagaaga gtacccagga
     3841 aacgcactgc atgaagcatg gagagaagct tcacctattc tgtgaggaag atgggcaggc
     3901 cctttgctgg gtgtgtgccc agtctgggaa acaccgggac cacaccaggg tccctattga
     3961 agaggctgct aaggtatacc aggagaagat ccacgtggct ttagaaaaac tgagaaaggg
     4021 gaaagagttg gccgagaaga tggaaatgga tctcacgatg caaagaacag actggaagag
     4081 gaacattgac acccagaagt cgaggattca cgcagagttc gcacttcaga atagcttgct
     4141 ggctcaggag gagcagaggc agctgcagag gctggagaag gatcaaaggg agtacctgag
     4201 actcctgggg aagaaggagg ctgagctggc tgagaagaac caggccctgc aggagctgat
     4261 ctcagagctg gagaggagga ttcgtggttc agagctggag ctactgcagg aggtgaggat
     4321 catcctggaa aggagtggat cctggaacct ggacacgtta gatattgacg ccccagacct
     4381 aacaagcaca tgccctgtgc cagggcggaa gaagatgctg aggacgtgtt gggttcatat
     4441 tactctggat cgcaacaccg ccaactcatg gctcatcatc tcaaaggatc ggagacaagt
     4501 gaggatggga gacacccatc agaacgtgtc tgacaataag gagaggttta gtaattaccc
     4561 catggtgcta ggtgcccaga gattctcctc tgggaagatg tactgggagg tagatgtgac
     4621 tcaaaaggag gcctgggatc tgggggtttg cagagattct gttcagagga aagggcagtt
     4681 ttcactcagt cccgagaatg gcttctggac catttggtta tggcaagaca gctatgaggc
     4741 tggtaccagt cctcagacca ccctccacat tcaagtacct ccatgccaaa ttgggatctt
     4801 tgtggactat gaggctggcg ttgtctcctt ctacaacata actgaccatg gctccctcat
     4861 ttacaccttc tcggagtgtg tttttgctgg acctctgcga cctttcttca atgttggttt
     4921 caattatagt gggggaaatg cagcgcctct aaagctctgt ccactaaaga tgtgagcggc
     4981 cgctctagat aaatgatagc ttgatctggt taccactaaa ccagcctcaa gaacacccga
     5041 atggagtctc taagctacat aataccaact tacactttac aaaatgttgt cccccaaaat
     5101 gtagccattc gtatctgctc ctaataaaaa gaaagtttct tcac
//
