LOCUS       sequence-448435-        3835 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid donor template for CRISPR/Cas9 mediated UGCG knockout by 
            EGFP insertion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3835)
  AUTHORS   Sato Y, Osada E, Ushiki T, Maeda T, Manome Y
  TITLE     UDP-glucose ceramide glucosyltransferase specifically upregulated in
            plasmacytoid dendritic cells regulates type I interferon production 
            upon CpG stimulation.
  JOURNAL   Biochem Biophys Res Commun. 2024 Sep 18;733:150703. doi: 
            10.1016/j.bbrc.2024.150703.
  PUBMED    39307111
REFERENCE   2  (bases 1 to 3835)
  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: "Biochem 
            Biophys Res Commun."; date: "2024-09-18"; volume: "733:150703. doi";
            pages: " 10.1016/j.bbrc.2024.150703"
COMMENT     Sequence Label: pUCmini-UGCG-EGFP
FEATURES             Location/Qualifiers
     source          1..3835
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        339..849
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     768..786
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     regulatory      865..874
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(871..873,874..876,877..1590)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(871..873,874..876,877..1587)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(916..937)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1177..1196)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1524..1545
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1596..1707
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2080..2668)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2160..2179)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2839..3630,3631..3699))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3462..3481
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3700..3804)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccaatgatct cgagtcacat ataaaataca ctaacaatag ctgatgagct ttaaaaaatc
       61 gcaaaaaatc tcataatgtt ttaagaaagt ttatgaattt gtgttggacc gcattcaaag
      121 ccatgcttac ctggttctac ttcccttcat gtttagatcc tcttttagaa agatgtcaag
      181 ttttaaaaat cttaacttag aatttaggga tttattgata gcttgtcagt gctttgattt
      241 gacactaatg tattctatag aaataatttt tatatcattt gctttttagc cgattacacg
      301 tcaacgcatg ctggggatgc ggtgggctga attcccacgg ggttggggtt gcgccttttc
      361 caaggcagcc ctgggtttgc gcagggacgc ggctgctctg ggcgtggttc cgggaaacgc
      421 agcggcgccg accctgggtc tcgcacattc ttcacgtccg ttcgcagcgt cacccggatc
      481 ttcgccgcta cccttgtggg ccccccggcg acgcttcctg ctccgcccct aagtcgggaa
      541 ggttccttgc ggttcgcggc gtgccggacg tgacaaacgg aagccgcacg tctcactagt
      601 accctcgcag acggacagcg ccagggagca atggcagcgc gccgaccgcg atgggctgtg
      661 gccaatagcg gctgctcagc ggggcgcgcc gagagcagcg gccgggaagg ggcggtgcgg
      721 gaggcggggt gtggggcggt agtgtgggcc ctgttcctgc ccgcgcggtg ttccgcattc
      781 tgcaagcctc cggagcgcac gtcggcagtc ggctccctcg ttgaccgaat caccgacctc
      841 tctccccagg gggatccacc ggtcgccacc atggtgagca agggcgagga gctgttcacc
      901 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
      961 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
     1021 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     1081 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     1141 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     1201 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     1261 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     1321 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     1381 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     1441 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa
     1501 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     1561 actctcggca tggacgagct gtacaagtaa gtcgactgtg ccttctagtt gccagccatc
     1621 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     1681 ttcctaataa aatgaggaaa ttgcatcatc gataagaagc caactgacaa acagccttat
     1741 agcaagctcc caggtgtctc tcttctgaaa ccactgaaag gggtagatcc taacttaatc
     1801 aacaacctgg aaacattctt tgaattggat tatcccaaag taagttcagt gttacggttt
     1861 tttgttttgt tttgttttgt tttgttcccc tcatgataaa catttaggga ttttaacact
     1921 gtataaggtg aaggtattaa aaattcatga tgatattggc ctcataaaat ataaccttta
     1981 gtagtagtca tcacagaaca gagctagcat cggaaagaac atgtgagcaa aaggccagca
     2041 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     2101 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     2161 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     2221 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     2281 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     2341 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     2401 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     2461 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     2521 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     2581 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     2641 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     2701 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     2761 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     2821 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     2881 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     2941 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     3001 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     3061 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     3121 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     3181 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     3241 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     3301 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     3361 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     3421 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     3481 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     3541 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     3601 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     3661 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     3721 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     3781 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
