LOCUS       sequence-338118-        3756 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Universal donor plasmid for CRISPitope method encoding mScarlet 
            fluorescent protein and blasticidin resistance cassette [p.M1G].
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3756)
  AUTHORS   Effern M, Glodde N, Braun M, Liebing J, Boll HN, Yong M, Bawden E, 
            Hinze D, van den Boorn-Konijnenberg D, Daoud M, Aymans P, Landsberg 
            J, Smyth MJ, Flatz L, Tuting T, Bald T, Gebhardt T, Holzel M
  TITLE     Adoptive T Cell Therapy Targeting Different Gene Products Reveals 
            Diverse and Context-Dependent Immune Evasion in Melanoma.
  JOURNAL   Immunity. 2020 Sep 15;53(3):564-580.e9. doi: 
            10.1016/j.immuni.2020.07.007. Epub 2020 Aug 3.
  PUBMED    32750334
REFERENCE   2  (bases 1 to 3756)
  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.immuni.2020.07.007"; journalName: "Immunity"; date: 
            "2020-09-15- 15"; volume: "53"; issue: "3"; pages: "564-580.e9"
COMMENT     Sequence Label: pCRISPaint-mScarlet-BlastR [M1G]
FEATURES             Location/Qualifiers
     source          1..3756
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..49
                     /note="synthetic polyadenylation signal"
     misc_feature    63..154
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     103..122
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     CDS             227..919
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="VSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGGA
                     VTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKM
                     ALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS
                     TGGMDELYK"
     primer_bind     complement(357..375)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     625..644
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             929..982
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     polyA_signal    1464..1585
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1475..1494)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1529..1548
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1833..1850)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2004..2592)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2084..2103)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2792..3583,3584..3652))
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctcttaatga tagctaggat cgctagcggg
      181 ccagtaccca aaaagcgggg ggtctggtgg cagtggaggg ggatccgtga gcaagggcga
      241 ggcagtgatc aaggagttca tgcggttcaa ggtgcacatg gagggctcca tgaacggcca
      301 cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa
      361 gctgaaggtg accaagggtg gccccctgcc cttctcctgg gacatcctgt cccctcagtt
      421 catgtacggc tccagggcct tcaccaagca ccccgccgac atccccgact actataagca
      481 gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggcgccgt
      541 gaccgtgacc caggacacct ccctggagga cggcaccctg atctacaagg tgaagctccg
      601 cggcaccaac ttccctcctg acggccccgt aatgcagaag aagacaatgg gctgggaagc
      661 gtccaccgag cggttgtacc ccgaggacgg cgtgctgaag ggcgacatta agatggccct
      721 gcgcctgaag gacggcggcc gctacctggc ggacttcaag accacctaca aggccaagaa
      781 gcccgtgcag atgcccggcg cctacaacgt cgaccgcaag ttggacatca cctcccacaa
      841 cgaggactac accgtggtgg aacagtacga acgctccgag ggccgccact ccaccggcgg
      901 catggacgag ctgtacaagg gatccggaga gggcagagga agtctgctaa catgcggtga
      961 cgtcgaggag aatcctggcc cagggaagac cttcaacatc tctcagcagg atctggagct
     1021 ggtggaggtc gccactgaga agatcaccat gctctatgag gacaacaagc accatgtcgg
     1081 ggcggccatc aggaccaaga ctggggagat catctctgct gtccacattg aggcctacat
     1141 tggcagggtc actgtctgtg ctgaagccat tgccattggg tctgctgtga gcaacgggca
     1201 gaaggacttt gacaccattg tggctgtcag gcacccctac tctgatgagg tggacagatc
     1261 catcagggtg gtcagcccct gtggcatgtg cagagagctc atctctgact atgctcctga
     1321 ctgctttgtg ctcattgaga tgaatggcaa gctggtcaaa accaccattg aggaactcat
     1381 ccccctcaag tacaccagga actaacacct ggtgcatgac ccgcaagccc ggtgcctgag
     1441 cggcccgctt cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag
     1501 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     1561 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt
     1621 tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat
     1681 cgataaggat ccgtcgaccg atgcccttga gagccttcaa cccagtcagc tccttccggt
     1741 gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc atgcaactcg
     1801 taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     1861 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     1921 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     1981 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2041 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2101 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2161 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     2221 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2281 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     2341 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     2401 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     2461 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     2521 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     2581 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     2641 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     2701 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     2761 agcggccgca aatgctaaac cactgcagtg gttaccagtg cttgatcagt gaggcaccga
     2821 tctcagcgat ctgcctattt cgttcgtcca tagtggcctg actccccgtc gtgtagatca
     2881 ctacgattcg tgagggctta ccatcaggcc ccagcgcagc aatgatgccg cgagagccgc
     2941 gttcaccggc ccccgatttg tcagcaatga accagccagc agggagggcc gagcgaagaa
     3001 gtggtcctgc tactttgtcc gcctccatcc agtctatgag ctgctgtcgt gatgctagag
     3061 taagaagttc gccagtgagt agtttccgaa gagttgtggc cattgctact ggcatcgtgg
     3121 tatcacgctc gtcgttcggt atggcttcgt tcaactctgg ttcccagcgg tcaagccggg
     3181 tcacatgatc acccatatta tgaagaaatg cagtcagctc cttagggcct ccgatcgttg
     3241 tcagaagtaa gttggccgcg gtgttgtcgc tcatggtaat ggcagcacta cacaattctc
     3301 ttaccgtcat gccatccgta agatgctttt ccgtgaccgg cgagtactca accaagtcgt
     3361 tttgtgagta gtgtatacgg cgaccaagct gctcttgccc ggcgtctata cgggacaaca
     3421 ccgcgccaca tagcagtact ttgaaagtgc tcatcatcgg gaatcgttct tcggggcgga
     3481 aagactcaag gatcttgccg ctattgagat ccagttcgat atagcccact cttgcaccca
     3541 gttgatcttc agcatctttt actttcacca gcgtttcggg gtgtgcaaaa acaggcaagc
     3601 aaaatgccgc aaagaaggga atgagtgcga cacgaaaatg ttggatgctc atactcgtcc
     3661 tttttcaata ttattgaagc atttatcaga gttactagta cgtctctcaa ggataagtaa
     3721 gtaatattaa ggtacgggag gtattggaca ggccgc
//
