ArticleNucleic acids research2012
Analysis of alternative splicing of cassette exons at single-cell level using two fluorescent proteins.
Article in Nucleic acids research, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed, 28 citations in OpenAlex.
- Functional analysis of theAnimal biotechnology · 2024Article
- High-throughput Flow Cytometry Assay to Investigate TDP43 Splicing Function.Bio-protocol · 2020Article
- Dissecting splicing decisions and cell-to-cell variability with designed sequence libraries.Nature communications · 2019Article
- Functional reassessment of PAX6 single nucleotide variants by in vitro splicing assay.European journal of human genetics : EJHG · 2019Article
- Heterozygous Mutations in TBX1 as a Cause of Isolated Hypoparathyroidism.The Journal of clinical endocrinology and metabolism · 2018Article
- Article
- Mapping Whole-Transcriptome Splicing in Mouse Hematopoietic Stem Cells.Stem cell reports · 2017Article
- Comparative study reveals better far-red fluorescent protein for whole body imaging.Scientific reports · 2015Article
- Method for quantitative analysis of nonsense-mediated mRNA decay at the single cell level.Scientific reports · 2015Article
- Single-cell transcriptomics reveals bimodality in expression and splicing in immune cells.Nature · 2013Article
Corrections and comments
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Authors and funding
7 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Alternative splicing plays a major role in increasing proteome complexity and regulating gene expression. Here, we developed a new fluorescent protein-based approach to quantitatively analyze the alternative splicing of a target cassette exon (skipping or inclusion), which results in an open-reading frame shift. A fragment of a gene of interest is cloned between red and green fluorescent protein (RFP and GFP)-encoding sequences in such a way that translation of the normally spliced full-length transcript results in expression of both RFP and GFP. In contrast, alternative exon skipping results in the synthesis of RFP only. Green and red fluorescence intensities can be used to estimate the proportions of normal and alternative transcripts in each cell. The new method was successfully tested for human PIG3 (p53-inducible gene 3) cassette exon 4. Expected pattern of alternative splicing of PIG3 minigene was observed, including previously characterized effects of UV light irradiation and specific mutations. Interestingly, we observed a broad distribution of normal to alternative transcript ratio in individual cells with at least two distinct populations with ∼45% and >95% alternative transcript. We believe that this method is useful for fluorescence-based quantitative analysis of alternative splicing of target genes in a variety of biological models.
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