LOCUS       sequence-192221-        4141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Clover from the EF1a promoter in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4141)
  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 4141)
  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-Clover
FEATURES             Location/Qualifiers
     source          1..4141
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(53..86)
                     /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(150..738)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(230..249)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1063..1857)
                     /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(1063..1857)
                     /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(1174..1193)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1784..1803
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     polyA_signal    2042..2266
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2255..2272
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     primer_bind     2605..2624
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    2623..2678
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     2724..2743
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     primer_bind     3440..3461
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3503..3512
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     LTR             3550..3818
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     promoter        complement(3831..4042)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
ORIGIN
        1 ccatatatgg gctatgaact aatgaccccg taattgatta ctattactta agataacttc
       61 gtatagcata cattatacga agttatgaac atgtgagcaa aaggccagca aaaggccagg
      121 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
      181 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
      241 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
      301 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
      361 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
      421 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
      481 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
      541 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
      601 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
      661 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
      721 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
      781 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
      841 atccttttaa attaaaaatg aagttttagc acgtgctatt attgaagcat ttatcagggt
      901 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
      961 ccgcgcacat ttccccgaaa agtgccacct gatgcggtgt gaaataccgc acagatgcgt
     1021 aaggagaaaa taccgcatca ggaaattgta agcgttaata attcagaaga actcgtcaag
     1081 aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa
     1141 gcggtcagcc cattcgccgc caagctcttc agcaatatca cgggtagcca acgctatgtc
     1201 ctgatagcgg tccgccaccc ccagccggcc acagtcgatg aatccagaaa agcggccatt
     1261 ttccaccatg atattcggca agcaggcatc gccatgggtc acgacgagat cctcgccgtc
     1321 gggcatgctc gccttgagcc tggcgaacag ttcggctggc gcgagcccct gatgctcttc
     1381 gtccagatca tcctgatcga caagaccggc ttccatccga gtacgtgctc gctcgatgcg
     1441 atgtttcgct tggtggtcga atgggcaggt agccggatca agcgtatgca gccgccgcat
     1501 tgcatcagcc atgatggata ctttctcggc aggagcaagg tgagatgaca ggagatcctg
     1561 ccccggcact tcgcccaata gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac
     1621 agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct cgtcttgcag
     1681 ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga
     1741 cagccggaac acggcggcat cagagcagcc gattgtctgt tgtgcccagt catagccgaa
     1801 tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg
     1861 aaacgatcct catcctgtct cttgatcaga gcttgatccc ctgcgccatc agatccttgg
     1921 cggcaagaaa gccatccagt ttactttgca gggcttccca accttaccag agggcgcccc
     1981 agctggcaat tccggttcgc ttgctgtcca taaaaccgcc cagtagaaga ggcgcgccaa
     2041 accatagagc ccaccgcatc cccagcatgc ctgctattgt cttcccaatc ctcccccttg
     2101 ctgtcctgcc ccaccccacc ccccagaata gaatgacacc tactcagaca atgcgatgca
     2161 atttcctcat tttattagga aaggacagtg ggagtggcac cttccagggt caaggaaggc
     2221 acgggggagg ggcaaacaac agatggctgg caactagaag gcacagtcga gggctagggg
     2281 accctcagtt agcctccccc atctcccgag aagagggaca gctatgactg ggagtagtca
     2341 ggagaggagg aaaaatctgg ctagtaaaac atgtaaggaa aattttaggg atgttaaaga
     2401 aaaaaataac acaaaacaaa atataaaaaa aatctaacct caagtcaagg cttttctatg
     2461 gaataaggaa tggacagcag ggggctgttt catatactga tgacctcttt atagccaacc
     2521 tttgttcatg gcagccagca tatgggcata tgttgccaaa ctctaaacca aatactcatt
     2581 ctgatgtttt aaatgatttg ccctcccata tgtccttccg agtgagagac acaaaaaatt
     2641 ccaacacact attgcaatga aaataaattt cctttattag ccagaagtca gatgctcaag
     2701 gggcttcatg atgtccccat aatttttggc agagggaaaa agatctcagt ggtatttgtg
     2761 agccagggta ttggccacac cagccaccac cttctgatag gcagcctgca cgcggccgcc
     2821 taggcggcgg ttacgaactc cagcaggacc atgtgatcgc gcttctcgtt ggggtctttg
     2881 ctcagggcgg actgatggct caggtagtgg ttgtcgggca gcagcacggg gccgtcgccg
     2941 atgggggtgt tctgctggta gtggtcggcg agctgcacgc tgccgtcctc aacgttgtgg
     3001 cggatcttga agttagcctt gatgccgttc ttctgcttgt cggccgtgat atagacgttg
     3061 tggctgttga agttgtactc cagcttgtgc cccaggatgt tgccgtcctc cttgaagtcg
     3121 atgcccttca gctcgatgcg gttcaccagg gtgtcgccct cgaacttcac ctcggcgcgg
     3181 gtcttgtagg taccgtcgtc cttgaaagag atggtgcgct cctggacgta gccttcgggc
     3241 atggcggact tgaagaagtc gtgctgcttc atgtggtcgg ggtagcggct gaagcaggcc
     3301 acgccgtagc cgaaggtggt cacgagggtg ggccagggca cgggcagctt gccggtggtg
     3361 cagatgaact tcagggtcag cttgccgttg gtggcatcgc cctcgccctc gccgcggacg
     3421 ctgaacttgt ggccgtttac gtcgccgtcc agctcgacca ggatgggcac caccccggtg
     3481 aacagctcct cgcccttgct caccatggtg gcgaccggta gcgctagcgt aggcgccggt
     3541 cacagcttgg atctgtaacg gcgcagaaca gaaaacgaaa caaagacgta gagttgagca
     3601 agcagggtca ggcaaagcgt ggagagccgg ctgagtctag gtaggctcca agggagcgcc
     3661 ggacaaaggc ccggtctcga cctgagcttt aaacttacct agacggcgga cgcagttcag
     3721 gaggcaccac aggcgggagg cggcagaacg cgactcaacc ggcgtggatg gcggcctcag
     3781 gtagggcggc gggcgcgtga aggagagatg cgagcccctc gaagcttcag ctgtgttctg
     3841 gcggcaaacc cgttgcgaaa aagaacgttc acggcgacta ctgcacttat atacggttct
     3901 cccccaccct cgggaaaaag gcggagccag tacacgacat cactttccca gtttaccccg
     3961 cgccaccttc tctaggcacc ggttcaattg ccgacccctc cccccaactt ctcggggact
     4021 gtgggcgatg tgcgctctgc ccactgacgg gcaccggagc gatcgcagat cgtcaatggg
     4081 cgggggtcgt tgggcggtca gccaggcggg ccatttaccg taagttatgt aacgcggaac
     4141 t
//
