Analysis of pollen-specific alternative splicing in Arabidopsis thaliana via semi-quantitative PCR

Search for this publication on Google Scholar
Loraine, A., Estrada, A., Freese, N. H., & Blakley, I. (2015). Analysis of pollen-specific alternative splicing in Arabidopsis thaliana via semi-quantitative PCR. Peerj. https://doi.org/doi:10.7717/peerj.919
Download PDF

Analytics

208 views ◎
36 downloads ⇓


Description

Faculty, Alternative splicing enables a single gene to produce multiple mRNA isoforms by varying splice site selection. In animals, alternative splicing of mRNA isoforms between cell types is widespread and supports cellular differentiation. In plants, at least 20% of multi-exon genes are alternatively spliced, but the extent and significance of tissue-specific splicing is less well understood, partly because it is difficult to isolate cells of a single type. Pollen is a useful model system to study tissue-specific splicing in higher plants because pollen grains contain only two cell types and can be collected in large amounts without damaging cells. Previously, we identified pollen-specific splicing patterns by comparing RNA-Seq data from Arabidopsis pollen and leaves. Here, we used semi-quantitative PCR to validate pollen-specific splicing patterns among genes where RNA-Seq data analysis indicated splicing was most different between pollen and leaves. PCR testing confirmed eight of nine alternative splicing patterns, and results from the ninth were inconclusive. In four genes, alternative transcriptional start sites coincided with alternative splicing. This study highlights the value of the low-cost PCR assay as a method of validating RNA-Seq results.

Details
Title

Analysis of pollen-specific alternative splicing in Arabidopsis thaliana via semi-quantitative PCR

Author(s)

Loraine, Ann
Estrada, April
Freese, Nowlan H.
Blakley, Ivory

UNC Charlotte Affiliate Type

Faculty

Extent

1 online resource

Date Created

2015

Genre

articles

Resource Type

text

Abstract

Alternative splicing enables a single gene to produce multiple mRNA isoforms by varying splice site selection. In animals, alternative splicing of mRNA isoforms between cell types is widespread and supports cellular differentiation. In plants, at least 20% of multi-exon genes are alternatively spliced, but the extent and significance of tissue-specific splicing is less well understood, partly because it is difficult to isolate cells of a single type. Pollen is a useful model system to study tissue-specific splicing in higher plants because pollen grains contain only two cell types and can be collected in large amounts without damaging cells. Previously, we identified pollen-specific splicing patterns by comparing RNA-Seq data from Arabidopsis pollen and leaves. Here, we used semi-quantitative PCR to validate pollen-specific splicing patterns among genes where RNA-Seq data analysis indicated splicing was most different between pollen and leaves. PCR testing confirmed eight of nine alternative splicing patterns, and results from the ninth were inconclusive. In four genes, alternative transcriptional start sites coincided with alternative splicing. This study highlights the value of the low-cost PCR assay as a method of validating RNA-Seq results.

Subjects--Topics

Bioinformatics
Pollen
Nucleic acids

Is Part Of

PeerJ

Language

English

Published Version (Please cite this version)

doi:10.7717/peerj.919

Permalink

http://hdl.handle.net/20.500.13093/work:743

Rights

This item may be protected by copyright and other related rights. Atkins Library provides access to this item for educational and research purposes only; other uses require the permission of the copyright holder.