LOCUS       Exported                4726 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Red fluorescent reporter for assessing CRISPR gRNA activity.
ACCESSION   .
VERSION     .
KEYWORDS    pGEmCherry2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4726)
  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 4726)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4726
                     /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"
     CDS             684..1391
                     /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(826..844)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1094..1113
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1514..1635
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1551..1570)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1605..1624
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1642..2097)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1779..1800)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1991..2010
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2124..2228
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2230..2587
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(2253..2273)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2438..2573
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2500..2519
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2622..3416
                     /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(2676..2695)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3286..3305
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3604..3623)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3648..3695
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4024..4612
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4513..4532
                     /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 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc gcgggcccgg
      661 gcgccaccat gtctagagga tccgtgagca agggcgagga ggataacctg gccatcatca
      721 aggagttcct gcgcttcaag gtgcacatgg agggctccgt gaacggccac gagttcgaga
      781 tcgagggcga gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga
      841 ccaagggtgg ccccctgccc ttcgcctggg acatcctgtc ccctcagttc atgtacggct
      901 ccaaggccta cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg
      961 agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc
     1021 aggactcctc cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact
     1081 tcccctccga cggccccgta atgcagaaga agaccatggg ctgggaggcc tcctccgagc
     1141 ggatgtaccc cgaggacggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg
     1201 acggcggcca ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc
     1261 tgcccggcgc ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca
     1321 ccatcgtgga acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc
     1381 tgtacaagta aagcggccgc gactctagat cataatcagc cataccacat ttgtagaggt
     1441 tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc
     1501 aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     1561 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     1621 catcaatgta tcttaaggcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
     1681 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     1741 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     1801 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     1861 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     1921 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     1981 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     2041 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag
     2101 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     2161 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     2221 ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa
     2281 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     2341 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     2401 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     2461 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     2521 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     2581 ttgcaaagat cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     2641 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     2701 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     2761 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc
     2821 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     2881 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     2941 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     3001 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     3061 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     3121 cagccgaact gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga
     3181 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     3241 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     3301 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     3361 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     3421 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     3481 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     3541 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct
     3601 aactgaaaca cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag
     3661 acagaataaa acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg
     3721 ctggcactct gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc
     3781 cttttcccca ccccatcccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg
     3841 ggcggcaggc cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact
     3901 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     3961 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     4021 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     4081 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     4141 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     4201 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     4261 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     4321 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     4381 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     4441 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4501 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4561 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4621 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     4681 tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcat
//
