LOCUS       sequence-307150-        3421 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 3421)
  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 3421)
  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-mScarletI. PGK-puro
FEATURES             Location/Qualifiers
     source          1..3421
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             829..1524
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(962..980)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1230..1249
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1557..1691
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1769..2268
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1804..1827)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2201..2220
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2280..2327
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2376..2975
                     /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(2376..2395)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2872..2892
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3004..3138
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(3232..3394,3395..3406)
                     /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 ggcgaggcag tgatcaagga gttcatgcgg ttcaaggtgc acatggaggg ctccatgaac
      901 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
      961 gccaagctga aggtgaccaa gggtggcccc ctgcccttct cctgggacat cctgtcccct
     1021 cagttcatgt acggctccag ggccttcatc aagcaccccg ccgacatccc cgactactat
     1081 aagcagtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
     1141 gccgtgaccg tgacccagga cacctccctg gaggacggca ccctgatcta caaggtgaag
     1201 ctccgcggca ccaacttccc tcctgacggc cccgtaatgc agaagaagac aatgggctgg
     1261 gaagcgtcca ccgagcggtt gtaccccgag gacggcgtgc tgaagggcga cattaagatg
     1321 gccctgcgcc tgaaggacgg cggacgctac ctggcggact tcaagaccac ctacaaggcc
     1381 aagaagcccg tgcagatgcc cggcgcctac aacgtggacc gcaagttgga catcacctcc
     1441 cacaacgagg actacaccgt ggtggaacag tacgaacgct ccgagggccg ccactccacc
     1501 ggcggcatgg acgagctgta caagtaatct agaactatag tgagtcgtat tacgtagatc
     1561 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     1621 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     1681 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     1741 gggaggtttt ttaattcgcg gccgcaccgg gtaggggagg cgcttttccc aaggcagtct
     1801 ggagcatgcg ctttagcagc cccgctgggc acttggcgct acacaagtgg cctctggcct
     1861 cgcacacatt ccacatccac cggtaggcgc caaccggctc cgttctttgg tggccccttc
     1921 gcgccacctt ctactcctcc cctagtcagg aagttccccc ccgccccgca gctcgcgtcg
     1981 tgcaggacgt gacaaatgga agtagcacgt ctcactagtc tcgtgcagat ggacagcacc
     2041 gctgagcaat ggaagcgggt aggcctttgg ggcagcggcc aatagcagct ttgctccttc
     2101 gctttctggg ctcagaggct gggaaggggt gggtccgggg gcgggctcag gggcgggctc
     2161 aggggcgggg cgggcgcccg aaggtcctcc ggaggcccgg cattctgcac gcttcaaaag
     2221 cgcacgtctg ccgcgctgtt ctcctcttcc tcatctccgg gcctttcgac ctgcagcctg
     2281 ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag gtgaggaact
     2341 aaaccgtcga cgatttcact agtctcgaag ccaccatgac cgagtacaag cccacggtgc
     2401 gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg
     2461 actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc
     2521 tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg
     2581 acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg
     2641 ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga
     2701 tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg
     2761 gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg
     2821 aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc
     2881 ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc
     2941 gcacctggtg catgacccgc aagcccggtg cctgactaga actatagtga gtcgtattac
     3001 gtagatccag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag
     3061 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     3121 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     3181 gaggtgtggg aggtttttta attaagctta aacaagaatc tctagttttc tttcttgctt
     3241 ttacttttac ttccttaata ctcaagtaca attttaatgg agtacttttt tacttttact
     3301 caagtaagat tctagccaga tacttttact tttaattgag taaaattttc cctaagtact
     3361 tgtactttca cttgagtaaa atttttgagt actttttaca cctctgctcg accatatggg
     3421 a
//
