LOCUS       sequence-223639-        4722 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Red fluorescent reporter for assessing CRISPR gRNA activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4722)
  AUTHORS   Hojland Knudsen C, Asgrimsdottir ES, Rahimi K, Gill KP, Frandsen S, 
            Hvolbol Buchholdt S, Chen M, Kjems J, Febbraro F, Denham M
  TITLE     A Modified Monomeric Red Fluorescent Protein Reporter for Assessing 
            CRISPR Activity.
  JOURNAL   Front Cell Dev Biol. 2018 May 15;6:54. doi: 
            10.3389/fcell.2018.00054. eCollection 2018.
  PUBMED    29868584
REFERENCE   2  (bases 1 to 4722)
  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: "Front Cell 
            Dev Biol."; date: "2018-05-15"; volume: "6:54. doi"; pages: " 
            10.3389/fcell.2018.00054. eCollection 2018"
COMMENT     Sequence Label: pGEmCherry2
FEATURES             Location/Qualifiers
     source          1..4722
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..566
                     /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"
     CDS             680..1387
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNLAIIKEFLRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(822..840)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1090..1109
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1510..1631
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1547..1566)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1601..1620
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1638..2093)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1775..1796)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1987..2006
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2120..2224
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2226..2583
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2226..2417
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2249..2269)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2434..2569
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2618..3412
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2618..3412
                     /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(2672..2691)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3282..3301
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3600..3619)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3644..3691
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4020..4608
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4509..4528
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acgtggaggt ctatataagc agagctggtt agtgaccgtc agatccgcta gcgctaccgg
      601 actcagatct cgagctcaag cttcgaattc tgcagtcgac ggtaccgcgg gcccgggcgc
      661 caccatgtct agaggatccg tgagcaaggg cgaggaggat aacctggcca tcatcaagga
      721 gttcctgcgc ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt tcgagatcga
      781 gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa
      841 gggtggcccc ctgcccttcg cctgggacat cctgtcccct cagttcatgt acggctccaa
      901 ggcctacgtg aagcaccccg ccgacatccc cgactacttg aagctgtcct tccccgaggg
      961 cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg tgacccagga
     1021 ctcctccctg caggacggcg agttcatcta caaggtgaag ctgcgcggca ccaacttccc
     1081 ctccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcct ccgagcggat
     1141 gtaccccgag gacggcgccc tgaagggcga gatcaagcag aggctgaagc tgaaggacgg
     1201 cggccactac gacgctgagg tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc
     1261 cggcgcctac aacgtcaaca tcaagttgga catcacctcc cacaacgagg actacaccat
     1321 cgtggaacag tacgaacgcg ccgagggccg ccactccacc ggcggcatgg acgagctgta
     1381 caagtaaagc ggccgcgact ctagatcata atcagccata ccacatttgt agaggtttta
     1441 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
     1501 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     1561 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     1621 aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     1681 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     1741 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     1801 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     1861 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     1921 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     1981 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     2041 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg
     2101 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     2161 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     2221 gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     2281 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     2341 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2401 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     2461 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     2521 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     2581 aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     2641 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     2701 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     2761 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc
     2821 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     2881 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     2941 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     3001 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     3061 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     3121 cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca
     3181 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     3241 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     3301 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     3361 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
     3421 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
     3481 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     3541 atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact
     3601 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     3661 aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg
     3721 cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     3781 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     3841 gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat
     3901 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     3961 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4021 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4081 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4141 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4201 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4261 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4321 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4381 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4441 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4501 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4561 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4621 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     4681 ttatcccctg attctgtgga taaccgtatt accgccatgc at
//
