Isolation and analysis of mRNAs from specific cell types of plants by ribosome immunopurification.
Isolation and analysis of mRNAs from specific cell types of plants by ribosome immunopurification.
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DOI：
10.1007/978-1-62703-221-6_19
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发表时间：
2013-01-01
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影响因子：
--
通讯作者：
Bailey-Serres, Julia
中科院分区：
文献类型：
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作者：
Mustroph, Angelika;Zanetti, M Eugenia;Bailey-Serres, Julia
Multiple ribosomes assemble onto an individual mRNA to form a polyribosome (polysome) complex. The epitope tagging of specific ribosomal proteins can enable the immunopurification of polysomes from crude cell extracts derived from cryopreserved tissue samples. Through expression of the epitope-tagged ribosomal protein in cell-type and regional specific domains of Arabidopsis thaliana and other organisms it is feasible to quantitatively assess the mRNAs that are associated with ribosomes with cell-specific resolution. Here we present detailed methods for development of transgenics that express a FLAG-tagged version of ribosomal protein L18 (RPL18) under the direction of individual promoters with specific domains of expression, the immunopurification of ribosomes, and bioinformatic analyses of the resultant datasets obtained by microarray profiling. This methodology provides researchers with the opportunity to assess rapid changes at the organ, tissue, regional or cell-type specific level of mRNAs that are associated with ribosomes and therefore engaged in translation.
