LOCUS       Exported                6083 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Expression of m-Cherry-tagged wildtype mouse Trim21 in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    pmCherry-C1-mTrim21
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6083)
  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 6083)
  AUTHORS   .
  TITLE     Direct Submission
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."
FEATURES             Location/Qualifiers
     source          1..6083
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             613..1320
                     /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(758..776)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1026..1045
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1357..2745
                     /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"
     polyA_signal    2871..2992
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2908..2927)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2962..2981
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2999..3454)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3136..3157)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3348..3367
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3481..3585
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3587..3944
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(3610..3630)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3795..3930
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3857..3876
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3979..4773
                     /codon_start=1
                     /gene="aph(3)-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4033..4052)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4643..4662
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4961..4980)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5005..5052
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5381..5969
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5870..5889
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"