Dataset Information

Whole-body scanning PCR; a highly sensitive method to study the biodistribution of mRNAs, noncoding RNAs and therapeutic oligonucleotides.

ABSTRACT: Efficient tissue-specific delivery is a crucial factor in the successful development of therapeutic oligonucleotides. Screening for novel delivery methods with unique tissue-homing properties requires a rapid, sensitive, flexible and unbiased technique able to visualize the in vivo biodistribution of these oligonucleotides. Here, we present whole body scanning PCR, a platform that relies on the local extraction of tissues from a mouse whole body section followed by the conversion of target-specific qPCR signals into an image. This platform was designed to be compatible with a novel RT-qPCR assay for the detection of siRNAs and with an assay suitable for the detection of heavily chemically modified oligonucleotides, which we termed Chemical-Ligation qPCR (CL-qPCR). In addition to this, the platform can also be used to investigate the global expression of endogenous mRNAs and non-coding RNAs. Incorporation of other detection systems, such as aptamers, could even further expand the use of this technology.

SUBMITTER: Boos JA

PROVIDER: S-EPMC3753639 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Publications

Whole-body scanning PCR; a highly sensitive method to study the biodistribution of mRNAs, noncoding RNAs and therapeutic oligonucleotides.

Boos Julien A JA Kirk David W DW Piccolotto Mari-Luz ML Zuercher Werner W Gfeller Sandro S Neuner Philippe P Dattler Andre A Wishart William L WL Von Arx Fabian F Beverly Michael M Christensen Jesper J Litherland Karine K van de Kerkhof Esther E Swart Pieter J PJ Faller Thomas T Beyerbach Armin A Morrissey David D Hunziker Juerg J Beuvink Iwan I

Nucleic acids research 20130613 15

Efficient tissue-specific delivery is a crucial factor in the successful development of therapeutic oligonucleotides. Screening for novel delivery methods with unique tissue-homing properties requires a rapid, sensitive, flexible and unbiased technique able to visualize the in vivo biodistribution of these oligonucleotides. Here, we present whole body scanning PCR, a platform that relies on the local extraction of tissues from a mouse whole body section followed by the conversion of target-speci ...[more]

PMID: 23766292

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Project description:Gene transcripts or mRNAs and long noncoding RNAs (lncRNAs) are differentially expressed during porcine skeletal muscle development. However, only a few studies have been conducted on skeletal muscle transcriptome in pigs based on timepoints according to the growth curve for porcine. Here, we investigated gene expression in Qingyu pigs at three different growth stages: the inflection point with the maximum growth rate (MGI), the inflection point of the gradually increasing stage to the rapidly increasing stage (GRI), and the inflection point of the rapidly increasing stage to the slowly increasing stage (RSI). Subsequently, we explored gene expression profiles during muscle development at the MGI, GRI and RSI stages by Ribo-Zero RNA sequencing. Qingyu pigs reached the MGI, GRI and RSI stages at 156.40, 23.82 and 288.97 days of age with 51.73, 3.14 and 107.03 kg body weight, respectively. A total of 14,530 mRNAs and 11,970 lncRNAs were identified at the three stages, and 645, 323 differentially expressed genes (DEGs) and 696, 760 differentially expressed lncRNAs (DELs) were identified in the GRI vs. MGI, and RSI vs. MGI, comparisons. Functional enrichment analysis revealed that genes involved in immune system development and energy metabolism (mainly relate to amino acid, carbohydrate and lipid) were enriched at the GRI and MGI stages, respectively, whereas genes involved in lipid metabolism were enriched at the RSI stage. We further characterized G1430, an abundant lncRNA. The full-length sequence (316 nt) of lncRNA G1430 was determined by rapid amplification of cDNA ends (RACE). Subcellular distribution analysis by quantitative real-time PCR (qRT-PCR) revealed that G1430 is a cytoplasmic lncRNA. Binding site prediction and dual luciferase assay showed that lncRNA G1430 directly binds to microRNA 133a (miR-133a). Our findings provide the basis for further investigation of the regulatory mechanisms and molecular genetics of muscle development in pigs.

Dataset Information

Whole-body scanning PCR; a highly sensitive method to study the biodistribution of mRNAs, noncoding RNAs and therapeutic oligonucleotides.

Publications

Whole-body scanning PCR; a highly sensitive method to study the biodistribution of mRNAs, noncoding RNAs and therapeutic oligonucleotides.

Similar Datasets

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