LOCUS       sequence-191171-        4252 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mCerulean from the EF1a promoter in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4252)
  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 4252)
  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-mCerulean
FEATURES             Location/Qualifiers
     source          1..4252
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    174..398
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     387..404
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     primer_bind     737..756
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    755..810
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     856..875
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     misc_feature    967..1032
                     /label=MCS
                     /note="multiple cloning site"
     CDS             complement(join(1033..1743,1744..1746,1747..1749))
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1075..1096)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1424..1443
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1683..1704
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1746..1755
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     LTR             1793..2061
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     promoter        complement(2074..2285)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    complement(2437..2470)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(2534..3122)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2614..2633)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3447..4241)
                     /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             complement(3447..4241)
                     /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(3558..3577)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4168..4187
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ctcatcctgt ctcttgatca gagcttgatc ccctgcgcca tcagatcctt ggcggcaaga
       61 aagccatcca gtttactttg cagggcttcc caaccttacc agagggcgcc ccagctggca
      121 attccggttc gcttgctgtc cataaaaccg cccagtagaa gaggcgcgcc aaaccataga
      181 gcccaccgca tccccagcat gcctgctatt gtcttcccaa tcctccccct tgctgtcctg
      241 ccccacccca ccccccagaa tagaatgaca cctactcaga caatgcgatg caatttcctc
      301 attttattag gaaaggacag tgggagtggc accttccagg gtcaaggaag gcacggggga
      361 ggggcaaaca acagatggct ggcaactaga aggcacagtc gagggctagg ggaccctcag
      421 ttagcctccc ccatctcccg agaagaggga cagctatgac tgggagtagt caggagagga
      481 ggaaaaatct ggctagtaaa acatgtaagg aaaattttag ggatgttaaa gaaaaaaata
      541 acacaaaaca aaatataaaa aaaatctaac ctcaagtcaa ggcttttcta tggaataagg
      601 aatggacagc agggggctgt ttcatatact gatgacctct ttatagccaa cctttgttca
      661 tggcagccag catatgggca tatgttgcca aactctaaac caaatactca ttctgatgtt
      721 ttaaatgatt tgccctccca tatgtccttc cgagtgagag acacaaaaaa ttccaacaca
      781 ctattgcaat gaaaataaat ttcctttatt agccagaagt cagatgctca aggggcttca
      841 tgatgtcccc ataatttttg gcagagggaa aaagatctca gtggtatttg tgagccaggg
      901 cattggccac accagccacc accttctgat aggcagcctg cacgcggccg cttatctaga
      961 tccggtggat cccgggcccg cggtaccgtc gactgcagaa ttcgaagctt gagctcgaga
     1021 tctgagtccg gacttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa
     1081 ctccagcagg accatgtgat cgcgcttctc gttggggtct ttgctcagct tggactgggt
     1141 gctcaggtag tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg
     1201 gtagtggtcg gcgagctgca cgctgccgtc ctcgatgttg tggcggatct tgaagttggc
     1261 cttgatgccg ttcttctgct tgtcggcggt gatatagacg ttgtcgctga tggcgttgta
     1321 ctccagcttg tgccccagga tgttgccgtc ctccttgaag tcgatgccct tcagctcgat
     1381 gcggttcacc agggtgtcgc cctcgaactt cacctcggcg cgggtcttgt agttgccgtc
     1441 gtccttgaag aagatggtgc gctcctggac gtagccttcg ggcatggcgg acttgaagaa
     1501 gtcgtgctgc ttcatgtggt cggggtagcg ggcgaagcac tgcacgcccc aggtcagggt
     1561 ggtcacgagg gtgggccagg gcacgggcag cttgccggtg gtgcagatga acttcagggt
     1621 cagcttgccg taggtggcat cgccctcgcc ctcgccggac acgctgaact tgtggccgtt
     1681 tacgtcgccg tccagctcga ccaggatggg caccaccccg gtgaacagct cctcgccctt
     1741 gctcaccatg gtggcgaccg gtagcgctag cgtaggcgcc ggtcacagct tggatctgta
     1801 acggcgcaga acagaaaacg aaacaaagac gtagagttga gcaagcaggg tcaggcaaag
     1861 cgtggagagc cggctgagtc taggtaggct ccaagggagc gccggacaaa ggcccggtct
     1921 cgacctgagc tttaaactta cctagacggc ggacgcagtt caggaggcac cacaggcggg
     1981 aggcggcaga acgcgactca accggcgtgg atggcggcct caggtagggc ggcgggcgcg
     2041 tgaaggagag atgcgagccc ctcgaagctt cagctgtgtt ctggcggcaa acccgttgcg
     2101 aaaaagaacg ttcacggcga ctactgcact tatatacggt tctcccccac cctcgggaaa
     2161 aaggcggagc cagtacacga catcactttc ccagtttacc ccgcgccacc ttctctaggc
     2221 accggttcaa ttgccgaccc ctccccccaa cttctcgggg actgtgggcg atgtgcgctc
     2281 tgcccactga cgggcaccgg agcgatcgca gatcgtcaat gggcgggggt cgttgggcgg
     2341 tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat atgggctatg
     2401 aactaatgac cccgtaattg attactatta cttaagataa cttcgtatag catacattat
     2461 acgaagttat gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     2521 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     2581 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     2641 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     2701 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     2761 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2821 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2881 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2941 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     3001 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3061 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3121 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3181 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3241 aatgaagttt tagcacgtgc tattattgaa gcatttatca gggttattgt ctcatgagcg
     3301 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     3361 gaaaagtgcc acctgatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     3421 atcaggaaat tgtaagcgtt aataattcag aagaactcgt caagaaggcg atagaaggcg
     3481 atgcgctgcg aatcgggagc ggcgataccg taaagcacga ggaagcggtc agcccattcg
     3541 ccgccaagct cttcagcaat atcacgggta gccaacgcta tgtcctgata gcggtccgcc
     3601 acccccagcc ggccacagtc gatgaatcca gaaaagcggc cattttccac catgatattc
     3661 ggcaagcagg catcgccatg ggtcacgacg agatcctcgc cgtcgggcat gctcgccttg
     3721 agcctggcga acagttcggc tggcgcgagc ccctgatgct cttcgtccag atcatcctga
     3781 tcgacaagac cggcttccat ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg
     3841 tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc gcattgcatc agccatgatg
     3901 gatactttct cggcaggagc aaggtgagat gacaggagat cctgccccgg cacttcgccc
     3961 aatagcagcc agtcccttcc cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg
     4021 cccgtcgtgg ccagccacga tagccgcgct gcctcgtctt gcagttcatt cagggcaccg
     4081 gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg ctgacagccg gaacacggcg
     4141 gcatcagagc agccgattgt ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa
     4201 gcggccggag aacctgcgtg caatccatct tgttcaatca tgcgaaacga tc
//
