LOCUS       sequence-328199-        2753 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector backbone to support a knock-in of Linker-Dendra2 at
            the C-terminus of a target locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2753)
  AUTHORS   Bollen Y, Hageman JH, van Leenen P, Derks LLM, Ponsioen B, Buissant 
            des Amorie JR, Verlaan-Klink I, van den Bos M, Terstappen LWMM, van 
            Boxtel R, Snippert HJG
  TITLE     Efficient and error-free fluorescent gene tagging in human organoids
            without double-strand DNA cleavage.
  JOURNAL   PLoS Biol. 2022 Jan 28;20(1):e3001527. doi: 
            10.1371/journal.pbio.3001527.
  PUBMED    35089911
REFERENCE   2  (bases 1 to 2753)
  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: "PLoS 
            Biol."; date: "2022-01-28"; volume: "20(1):e3001527. doi"; pages: " 
            10.1371/journal.pbio.3001527"
COMMENT     Sequence Label: TVBB C-term-Dendra2
FEATURES             Location/Qualifiers
     source          1..2753
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(46..837,838..906))
                     /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     669..688
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(907..1011)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1139..1831
                     /codon_start=1
                     /product="monomeric photoswitchable green-to-red 
                     fluorescent protein from Dendronephthya"
                     /label=Dendra2
                     /note="mammalian codon-optimized"
                     /translation="MNTPGINLIKEDMRVKVHMEGNVNGHAFVIEGEGKGKPYEGTQTA
                     NLTVKEGAPLPFSYDILTTAVHYGNRVFTKYPEDIPDYFKQSFPEGYSWERTMTFEDKG
                     ICTIRSDISLEGDCFFQNVRFKGTNFPPNGPVMQKKTLKWEPSTEKLHVRDGLLVGNIN
                     MALLLEGGGHYLCDFKTTYKAKKVVQLPDAHFVDHRIEILGNDSDYNKVKLYEHAVARY
                     SPLPSQVW"
     primer_bind     complement(1869..1886)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2040..2628)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2120..2139)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
       61 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
      121 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
      181 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
      241 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
      301 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
      361 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
      421 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
      481 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
      541 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
      601 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
      661 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
      721 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
      781 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
      841 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
      901 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
      961 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     1021 ccgaaaagtg ccacctgatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     1081 gctacggaag agcgcgatcg cgtttaaagg ctcttcaggc ggaggcggca gcgccagcat
     1141 gaacaccccg ggaattaacc tgatcaagga ggacatgcgc gtgaaggtgc acatggaggg
     1201 caacgtgaac ggccacgcct tcgtgatcga gggcgagggc aagggcaagc cctacgaggg
     1261 cacccagacc gccaacctga ccgtgaagga gggcgccccc ctgcccttca gctacgacat
     1321 cctgaccacc gccgtgcact acggcaaccg ggtgttcacc aagtaccccg aggacatccc
     1381 cgactacttc aagcagagct tccccgaggg ctacagctgg gagcgcacca tgaccttcga
     1441 ggacaagggc atctgcacca tccgcagcga catcagcctg gagggcgact gcttcttcca
     1501 gaacgtgcgc ttcaagggca ccaacttccc ccccaacggc cccgtgatgc agaagaagac
     1561 cctgaagtgg gagcccagca ccgagaagct gcacgtgcgc gacggcctgc tggtgggcaa
     1621 catcaacatg gccctgctgc tggagggcgg cggccactac ctgtgcgact tcaagaccac
     1681 ctacaaggcc aagaaggtgg tgcagctgcc cgacgcccac ttcgtggacc accgcatcga
     1741 gatcctgggc aacgacagcg actacaacaa ggtgaagctg tacgagcacg ccgtggcccg
     1801 ctacagcccc ctgcccagcc aggtgtggtg acgaagagcg tttaaacgcg aaccggctct
     1861 tcgaattcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     1921 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     1981 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     2041 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     2101 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     2161 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     2221 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     2281 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     2341 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     2401 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     2461 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     2521 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     2581 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     2641 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     2701 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aag
//
