LOCUS       sequence-386921-        3109 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  To establish Ngn3ER cell line for endocrine differentiation 
            (4-hydroxytamoxifen activation, Puromycin resistant).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 3109)
  AUTHORS   Huang X, Gu W, Zhang J, Lan Y, Colarusso JL, Li S, Pertl C, Lu J, 
            Kim H, Zhu J, Breault DT, Sevigny J, Zhou Q
  TITLE     Stomach-derived human insulin-secreting organoids restore glucose 
            homeostasis.
  JOURNAL   Nat Cell Biol. 2023 Apr 27. doi: 10.1038/s41556-023-01130-y.
  PUBMED    37106062
REFERENCE   2  (bases 1 to 3109)
  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: "Nat Cell 
            Biol. 2023 Apr 27. doi: 10.1038/s41556-023-01130-y."
COMMENT     Sequence Label: pLenti-EF1a-Ngn3ER-PuroR-mCherry
FEATURES             Location/Qualifiers
     source          1..3109
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     protein_bind    1..25
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      53..62
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1673..1729
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1736..2332
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1736..1755)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2232..2252
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             2339..2392
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2399..3109
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2544..2562)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2812..2831
                     /label=mCherry-F
                     /note="mCherry, forward primer"
ORIGIN
        1 acaagtttgt acaaaaaagc aggctggcgc cggaaccaat tcagtcgacg ccgccaccat
       61 ggcgcctcat cccttggatg cgctcaccat ccaagtgtcc ccagagacac aacaaccttt
      121 tcccggagcc tcggaccacg aagtgctcag ttccaattcc accccaccta gccccactct
      181 catacctagg gactgctccg aagcagaagt gggtgactgc cgagggacct cgaggaagct
      241 ccgcgcccga cgcggagggc gcaacaggcc caagagcgag ttggcactca gcaaacagcg
      301 aagaagccgg cgcaagaagg ccaatgatcg ggagcgcaat cgcatgcaca acctcaactc
      361 ggcgctggat gcgctgcgcg gtgtcctgcc caccttcccg gatgacgcca aacttacaaa
      421 gatcgagacc ctgcgcttcg cccacaacta catctgggca ctgactcaga cgctgcgcat
      481 agcggaccac agcttctatg gcccggagcc ccctgtgccc tgtggagagc tggggagccc
      541 cggaggtggc tccaacgggg actggggctc tatctactcc ccagtctccc aagcgggtaa
      601 cctgagcccc acggcctcat tggaggaatt ccctggcctg caggtgccca gctccccatc
      661 ctatctgctc ccgggagcac tggtgttctc agacttcttg agggatccac gaaatgaaat
      721 gggtgcttca ggagacatga gggctgccaa cctttggcca agccctcttg tgattaagca
      781 cactaagaag aatagccctg ccttgtcctt gacagctgac cagatggtca gtgccttgtt
      841 ggatgctgaa ccgcccatga tctattctga atatgatcct tctagaccct tcagtgaagc
      901 ctcaatgatg ggcttattga ccaacctagc agatagggag ctggttcata tgatcaactg
      961 ggcaaagaga gtgccaggct ttggggactt gaatctccat gatcaggtcc accttctcga
     1021 gtgtgcctgg ctggagattc tgatgattgg tctcgtctgg cgctccatgg aacacccggg
     1081 gaagctcctg tttgctccta acttgctcct ggacaggaat caaggtaaat gtgtggaagg
     1141 catggtggag atctttgaca tgttgcttgc tacgtcaagt cggttccgca tgatgaacct
     1201 gcagggtgaa gagtttgtgt gcctcaaatc catcattttg cttaattccg gagtgtacac
     1261 gtttctgtcc agcaccttga agtctctgga agagaaggac cacatccacc gtgtcctgga
     1321 caagatcaca gacactttga tccacctgat ggccaaagct ggcctgactc tgcagcagca
     1381 gcatcgccgc ctagctcagc tccttctcat tctttcccat atccggcata tgagtaacaa
     1441 acgcatggag catctctaca acatgaaatg caagaacgtg gtacccctct atgacctgct
     1501 cctggagatg ttggatgccc accgccttca tgccccagcc agtcgcatgg gagtgccccc
     1561 agaggagccc agccagaccc agctggccac caccagctcc acttcagcac attccttaca
     1621 aacctactac atacccccgg aagcagaggg cttccccaac acgatcacta gtgccacgaa
     1681 cttctctctg ttaaagcaag caggagatgt tgaagaaaac cccgggcctg gatccatgac
     1741 cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac
     1801 cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgatc cggaccgcca
     1861 catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg
     1921 caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt
     1981 cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg
     2041 gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc
     2101 gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc
     2161 cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct tcctggagac
     2221 ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca ccgccgacgt
     2281 cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg ccggatccga
     2341 gggcagagga agtcttctaa catgcggtga cgtggaggag aatcccggcc ctatcgatat
     2401 ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc gcttcaaggt
     2461 gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg
     2521 cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt
     2581 cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg tgaagcaccc
     2641 cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt
     2701 gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg
     2761 cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat
     2821 gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg aggacggcgc
     2881 cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact acgacgctga
     2941 ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa
     3001 catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac agtacgaacg
     3061 cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtaa
//
