LOCUS       sequence-307155-        3466 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Fluorescent reporter for intestinal stem cells through ASCL2 
            transcriptional activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 3466)
  AUTHORS   Heinz MC, Oost KC, Snippert HJG
  TITLE     Introducing the Stem Cell ASCL2 Reporter STAR into Intestinal 
            Organoids.
  JOURNAL   STAR Protoc. 2020 Oct 8;1(3):100126. doi: 
            10.1016/j.xpro.2020.100126. eCollection 2020 Dec 18.
  PUBMED    33377020
REFERENCE   2  (bases 1 to 3466)
  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: "STAR 
            Protoc."; date: "2020-10-8"; volume: "1(3):100126. doi"; pages: " 
            10.1016/j.xpro.2020.100126. eCollection 2020 Dec 18"
COMMENT     Sequence Label: Tol2 8xSTAR-mNeonGreen-NLS. PGK-puro
FEATURES             Location/Qualifiers
     source          1..3466
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             829..1536
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      1084..1093
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1543..1563
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    1602..1736
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1814..2313
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1849..1872)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2246..2265
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2325..2372
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2421..3020
                     /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(2421..2440)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2917..2937
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3049..3183
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(3277..3439,3440..3451)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
ORIGIN
        1 agaggtgtaa agtacttgag taattttact tgattactgt acttaagtat tatttttggg
       61 gatttttact ttacttgagt acaattaaaa atcaatactt ttacttttac ttaattacat
      121 ttttttagaa aaaaaagtac tttttactcc ttacaatttt atttacagtc aaaaagtact
      181 tattttttgg agatcacttc attctatttt cccttgctat taccaaacca attgaattgc
      241 gctgatgccc agtttaattt aaatagatct atcgatcggt acccctaggg tgaaaaggca
      301 ggtgcaaagc aaatcctatc ccagcttcta gctaggtgaa aaggcaggtg caaagcaaat
      361 cctatcccag cttctagcta gggtgaaaag gcaggtgcaa agcaaatcct atcccagctt
      421 ctagctaggt gaaaaggcag gtgcaaagca aatcctatcc cagcttctag ctagggtgaa
      481 aaggcaggtg caaagcaaat cctatcccag cttctagcta ggtgaaaagg caggtgcaaa
      541 gcaaatccta tcccagcttc tagctagggt gaaaaggcag gtgcaaagca aatcctatcc
      601 cagcttctag ctaggtgaaa aggcaggtgc aaagcaaatc ctatcccagc ttctagctag
      661 cggcgcgccg gatccggagg ggacaagtgg agggaaagtg gggccgggcg gggggtgcct
      721 gggaagccag gctgcgctga cgtcactggg cgcgcaattc gggctggagc gctttaaaaa
      781 acgagcgtgc aagcagagag gctgctccac accgctcagg ccgcgaccat ggtgagcaag
      841 ggcgaggagg ataacatggc ctctctccca gcgacacatg agttacacat ctttggctcc
      901 atcaacggtg tggactttga catggtgggt cagggcaccg gcaatccaaa tgatggttat
      961 gaggagttaa acctgaagtc caccaagggt gacctccagt tctccccctg gattctggtc
     1021 cctcatatcg ggtatggctt ccatcagtac ctgccctacc ctgacgggat gtcgcctttc
     1081 caggccgcca tggtagatgg ctccggatac caagtccatc gcacaatgca gtttgaagat
     1141 ggtgcctccc ttactgttaa ctaccgctac acctacgagg gaagccacat caaaggagag
     1201 gcccaggtga aggggactgg tttccctgct gacggtcctg tgatgaccaa ctcgctgacc
     1261 gctgcggact ggtgcaggtc gaagaagact taccccaacg acaaaaccat catcagtacc
     1321 tttaagtgga gttacaccac tggaaatggc aagcgctacc ggagcactgc gcggaccacc
     1381 tacacctttg ccaagccaat ggcggctaac tatctgaaga accagccgat gtacgtgttc
     1441 cgtaagacgg agctcaagca ctccaagacc gagctcaact tcaaggagtg gcaaaaggcc
     1501 tttaccgatg tgatgggcat ggacgagctg tacaagtcca ggccaaaaaa gaaaagaaag
     1561 gtcatcgatt agtctagaac tatagtgagt cgtattacgt agatccagac atgataagat
     1621 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     1681 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     1741 acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaat
     1801 tcgcggccgc accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta
     1861 gcagccccgc tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca
     1921 tccaccggta ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact
     1981 cctcccctag tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa
     2041 atggaagtag cacgtctcac tagtctcgtg cagatggaca gcaccgctga gcaatggaag
     2101 cgggtaggcc tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag
     2161 aggctgggaa ggggtgggtc cgggggcggg ctcaggggcg ggctcagggg cggggcgggc
     2221 gcccgaaggt cctccggagg cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg
     2281 ctgttctcct cttcctcatc tccgggcctt tcgacctgca gcctgttgac aattaatcat
     2341 cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc gtcgacgatt
     2401 tcactagtct cgaagccacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     2461 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     2521 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     2581 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     2641 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     2701 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     2761 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     2821 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     2881 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     2941 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     3001 cccgcaagcc cggtgcctga ctagaactat agtgagtcgt attacgtaga tccagacatg
     3061 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     3121 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     3181 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt
     3241 ttttaattaa gcttaaacaa gaatctctag ttttctttct tgcttttact tttacttcct
     3301 taatactcaa gtacaatttt aatggagtac ttttttactt ttactcaagt aagattctag
     3361 ccagatactt ttacttttaa ttgagtaaaa ttttccctaa gtacttgtac tttcacttga
     3421 gtaaaatttt tgagtacttt ttacacctct gctcgaccat atggga
//
