LOCUS       sequence-192220-        4168 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mRuby2 from the EF1a promoter in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4168)
  AUTHORS   Geisinger JM, Turan S, Hernandez S, Spector LP, Calos MP
  TITLE     In vivo blunt-end cloning through CRISPR/Cas9-facilitated 
            non-homologous end-joining.
  JOURNAL   Nucleic Acids Res. 2016 Jan 13. pii: gkv1542.
  PUBMED    26762978
REFERENCE   2  (bases 1 to 4168)
  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: "Nucleic 
            Acids Res. 2016 Jan 13. pii: gkv1542."
COMMENT     Sequence Label: pKER-mRuby2
FEATURES             Location/Qualifiers
     source          1..4168
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(251..268)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    257..481
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             666..1460
                     /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             666..1460
                     /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(720..739)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1330..1349
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      1785..2373
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2274..2293
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    2437..2470
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2622..2833
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             2846..3114
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     regulatory      3152..3161
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3158..3868
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     primer_bind     complement(3948..3967)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4013..4068
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4067..4086)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
ORIGIN
        1 aaggttggct ataaagaggt catcagtata tgaaacagcc ccctgctgtc cattccttat
       61 tccatagaaa agccttgact tgaggttaga ttttttttat attttgtttt gtgttatttt
      121 tttctttaac atccctaaaa ttttccttac atgttttact agccagattt ttcctcctct
      181 cctgactact cccagtcata gctgtccctc ttctcgggag atgggggagg ctaactgagg
      241 gtcccctagc cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
      301 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
      361 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
      421 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
      481 gtttggcgcg cctcttctac tgggcggttt tatggacagc aagcgaaccg gaattgccag
      541 ctggggcgcc ctctggtaag gttgggaagc cctgcaaagt aaactggatg gctttcttgc
      601 cgccaaggat ctgatggcgc aggggatcaa gctctgatca agagacagga tgaggatcgt
      661 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
      721 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
      781 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
      841 aactgcaaga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
      901 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
      961 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     1021 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     1081 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     1141 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgagcatgc
     1201 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     1261 aaaatggccg cttttctgga ttcatcgact gtggccggct gggggtggcg gaccgctatc
     1321 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     1381 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     1441 ttcttgacga gttcttctga attattaacg cttacaattt cctgatgcgg tattttctcc
     1501 ttacgcatct gtgcggtatt tcacaccgca tcaggtggca cttttcgggg aaatgtgcgc
     1561 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1621 taaccctgat aaatgcttca ataatagcac gtgctaaaac ttcattttta atttaaaagg
     1681 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     1741 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     1801 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     1861 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     1921 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     1981 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     2041 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     2101 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     2161 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     2221 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     2281 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     2341 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     2401 ttcctggcct tttgctggcc ttttgctcac atgttcataa cttcgtataa tgtatgctat
     2461 acgaagttat cttaagtaat agtaatcaat tacggggtca ttagttcata gcccatatat
     2521 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     2581 ccgcccattg acgatctgcg atcgctccgg tgcccgtcag tgggcagagc gcacatcgcc
     2641 cacagtcccc gagaagttgg ggggaggggt cggcaattga accggtgcct agagaaggtg
     2701 gcgcggggta aactgggaaa gtgatgtcgt gtactggctc cgcctttttc ccgagggtgg
     2761 gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca acgggtttgc
     2821 cgccagaaca cagctgaagc ttcgaggggc tcgcatctct ccttcacgcg cccgccgccc
     2881 tacctgaggc cgccatccac gccggttgag tcgcgttctg ccgcctcccg cctgtggtgc
     2941 ctcctgaact gcgtccgccg tctaggtaag tttaaagctc aggtcgagac cgggcctttg
     3001 tccggcgctc ccttggagcc tacctagact cagccggctc tccacgcttt gcctgaccct
     3061 gcttgctcaa ctctacgtct ttgtttcgtt ttctgttctg cgccgttaca gatccaagct
     3121 gtgaccggcg cctacgctag cgctaccggt cgccaccatg gtgtctaagg gcgaagagct
     3181 gatcaaggaa aatatgcgta tgaaggtggt catggaaggt tcggtcaacg gccaccaatt
     3241 caaatgcaca ggtgaaggag aaggcaatcc gtacatggga actcaaacca tgaggatcaa
     3301 agtcatcgag ggaggacccc tgccatttgc ctttgacatt cttgccacgt cgttcatgta
     3361 tggcagccgt acttttatca agtacccgaa aggcattcct gatttcttta aacagtcctt
     3421 tcctgagggt tttacttggg aaagagttac gagatacgaa gatggtggag tcgtcaccgt
     3481 catgcaggac accagccttg aggatggctg tctcgtttac cacgtccaag tcagaggggt
     3541 aaactttccc tccaatggtc ccgtgatgca gaagaagacc aagggttggg agcctaatac
     3601 agagatgatg tatccagcag atggtggtct gaggggatac actcatatgg cactgaaagt
     3661 tgatggtggt ggccatctgt cttgctcttt cgtaacaact tacaggtcaa aaaagaccgt
     3721 cgggaacatc aagatgcccg gtatccatgc cgttgatcac cgcctggaaa ggttagagga
     3781 aagtgacaat gaaatgttcg tagtacaacg cgaacacgca gttgccaagt tcgccgggct
     3841 tggtggtggg atggacgagc tgtacaagta ggcggccgcg tgcaggctgc ctatcagaag
     3901 gtggtggctg gtgtggccaa tgccctggct cacaaatacc actgagatct ttttccctct
     3961 gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg ctaataaagg
     4021 aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact cggaaggaca
     4081 tatgggaggg caaatcattt aaaacatcag aatgagtatt tggtttagag tttggcaaca
     4141 tatgcccata tgctggctgc catgaaca
//
