pGEMHE-mCherry-mTrim21

LOCUS      pGEMHE-mCherry-mTrim21                5197 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  T7 promotor drives in vitro transcription of mCherry-tagged wildtype
            mouse TRIM21 mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    pGEMHE-mCherry-mTrim21
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5197)
  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 5197)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..5197
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        complement(67..171)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     239..257
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(295..317)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     417..436
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     630..652
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     644..661
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     645..661
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     668..687
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        668..686
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     5'UTR           718..760
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     regulatory      794..803
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             800..1507
                     /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(945..963)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1213..1232
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1544..2932
                     /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           2976..3101
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     promoter        complement(3206..3224)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3207..3224)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3242..3258)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3242..3258)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3255..3277)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3266..3282
                     /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(3290..3320)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3335..3356
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3473..3490)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3644..4232)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3724..3743)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4403..66)
                     /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     5026..5045
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
       61 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
      121 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
      181 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
      241 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
      301 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
      361 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
      421 atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt
      481 aaggagaaaa taccgcatca ggcgccattc gccattcagg ctgcgcaact gttgggaagg
      541 gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag
      601 gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag
      661 tgaattgtaa tacgactcac tatagggcga attaattcga gctcggtacc cagcttgctt
      721 gttctttttg cagaagctca gaataaacgc tcaactttgg cagatcaatt ccccggtgct
      781 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa
      841 ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat
      901 cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac
      961 caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc
     1021 caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga
     1081 gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca
     1141 ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt
     1201 cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg
     1261 gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga
     1321 cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct
     1381 gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac
     1441 catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct
     1501 gtacaagtcc ggactcagat ctcgagctca agcttcgaat tcaatgtctc tggaaaagat
     1561 gtgggaggag gtcacctgtt ctatctgcct ggatcccatg gtggagccta tgagtatcga
     1621 atgtggccat tgcttttgca aggaatgcat ttttgaagtt gggaagaatg ggggcagttc
     1681 atgtcccgag tgccggcaac agtttctgct ccgaaacctc aggcccaata gacatatagc
     1741 caacatggtg gaaaacctta aacagatagc ccagaatacc aagaagagta cccaggaaac
     1801 gcactgcatg aagcatggag agaagcttca cctattctgt gaggaagatg ggcaggccct
     1861 ttgctgggtg tgtgcccagt ctgggaaaca ccgggaccac accagggtcc ctattgaaga
     1921 ggctgctaag gtataccagg agaagatcca cgtggcttta gaaaaactga gaaaggggaa
     1981 agagttggcc gagaagatgg aaatggatct cacgatgcaa agaacagact ggaagaggaa
     2041 cattgacacc cagaagtcga ggattcacgc agagttcgca cttcagaata gcttgctggc
     2101 tcaggaggag cagaggcagc tgcagaggct ggagaaggat caaagggagt acctgagact
     2161 cctggggaag aaggaggctg agctggctga gaagaaccag gccctgcagg agctgatctc
     2221 agagctggag aggaggattc gtggttcaga gctggagcta ctgcaggagg tgaggatcat
     2281 cctggaaagg agtggatcct ggaacctgga cacgttagat attgacgccc cagacctaac
     2341 aagcacatgc cctgtgccag ggcggaagaa gatgctgagg acgtgttggg ttcatattac
     2401 tctggatcgc aacaccgcca actcatggct catcatctca aaggatcgga gacaagtgag
     2461 gatgggagac acccatcaga acgtgtctga caataaggag aggtttagta attaccccat
     2521 ggtgctaggt gcccagagat tctcctctgg gaagatgtac tgggaggtag atgtgactca
     2581 aaaggaggcc tgggatctgg gggtttgcag agattctgtt cagaggaaag ggcagttttc
     2641 actcagtccc gagaatggct tctggaccat ttggttatgg caagacagct atgaggctgg
     2701 taccagtcct cagaccaccc tccacattca agtacctcca tgccaaattg ggatctttgt
     2761 ggactatgag gctggcgttg tctccttcta caacataact gaccatggct ccctcattta
     2821 caccttctcg gagtgtgttt ttgctggacc tctgcgacct ttcttcaatg ttggtttcaa
     2881 ttatagtggg ggaaatgcag cgcctctaaa gctctgtcca ctaaagatgt gagcggccgc
     2941 tctagataaa tgatagcttg atctggttac cactaaacca gcctcaagaa cacccgaatg
     3001 gagtctctaa gctacataat accaacttac actttacaaa atgttgtccc ccaaaatgta
     3061 gccattcgta tctgctccta ataaaaagaa agtttcttca cattctaaaa aaaaaaaaaa
     3121 aaaaaaaaaa aaaaaaaaaa cccccccccc ccccccctgc aggcgcgccg ccggcgattt
     3181 aaattaatta atctagcttg agtattctat agtgtcacct aaatagcttg gcgtaatcat
     3241 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     3301 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     3361 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     3421 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     3481 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     3541 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     3601 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3661 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     3721 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     3781 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     3841 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     3901 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3961 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     4021 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     4081 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     4141 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     4201 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     4261 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     4321 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     4381 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     4441 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     4501 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     4561 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     4621 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     4681 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     4741 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     4801 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     4861 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     4921 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     4981 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     5041 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     5101 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     5161 cagcatcttt tactttcacc agcgtttctg ggtgagc
//