Extended Data Fig. 3: Characterization of minigene variants of MAPT with integrated dual-luciferase EXSISERS constructs.
From: Non-invasive and high-throughput interrogation of exon-specific isoform expression
a, A MAPT 4R-minigene was created by amplifying exon-adjacent intronic sequences between exon 9 and 10, and between 10 and 11. b, Immunoblot analysis (anti-pan-tau) of HEK293T cells, transiently transfected with a Pgk1 promoter-driven plasmid overexpressing MAPT 0N3R CDS yield a single 0N3R protein species and minor tau degradation products. c, Immunoblot analysis (anti-pan-tau) of HEK293T cells transfected with a minigene described in a) and its corresponding densitometric analysis (dephosphorylated lysate). Please note that it is challenging to obtain sufficiently separated bands for tau isoforms to perform precise densitometric quantification. Data shown in b and c represent 2 independent experiments. d, Overexposed and contrast-enhanced images of c) showed minor amounts of tau breakdown products and unspliced variants of the overexpressed luciferases. e, Fractional inclusion of exon 10 in HEK293T cells transfected with the minigene constructs described in a. By applying the conversion for the relative luminescence brightness of FLuc and NLuc (Extended Data Fig. 2c), the fractional inclusion of exon 10 for the WT-minigene variant is ~19% an 35% for the IVS10+16 C>T minigene. Data shown represent the mean ± s.d (n = 3 biological replicates), **** denotes a p-value smaller than 0.0001 in a two-tailed unpaired t-test (full statistical results are available in Supplementary Table 1). Unprocessed blots and numerical source data are provided in Source Data Extended Data Fig. 3.
