Closure of the Entire Fistula With Highly Effective Chemocauterization Using the Distal Hood Endoscope: A Novel Procedure for the Treatment of Pyriform Sinus Fistula.

Background: Pyriform sinus fistula (PSF) causes a recurrent abscess in the neck. Endoscopic chemocauterization with trichloroacetic acid (TCA) for PSF is a simple, reproducible, and reliable procedure for treating PSF; however, there is concern about complications caused by TCA overflowing into the larynx. To prevent these complications, we devised a highly effective chemocauterization using a distal hooded endoscope (HuDHE).

Laser Microdissection-Mediated Isolation and In Vitro Transcriptional Amplification of Plant RNA.

Laser microdissection of cells allows for isolation of specific cells of interest for downstream analyses including transcriptional profiling. Plant cells present unique challenges for laser microdissection due to their cellulosic cell walls and large vacuoles. Here we present protocols for plant tissue preparation, laser microdissection of select plant cells, and linear amplification of RNA from dissected cells.

DNA extraction from freeze-dried plant tissue with CTAB in a 96-well format.

This protocol is a modified version of DNA isolation using cetyltrimethylammonium bromide (CTAB) and 96-well plates. It is high-throughput, which facilitates the analysis of large mapping populations. The DNA yield is adequate for at least 100-500 polymerase chain reaction (PCR) procedures.

Loss of RNA-dependent RNA polymerase 2 (RDR2) function causes widespread and unexpected changes in the expression of transposons, genes, and 24-nt small RNAs.

Transposable elements (TEs) comprise a substantial portion of many eukaryotic genomes and are typically transcriptionally silenced. RNA-dependent RNA polymerase 2 (RDR2) is a component of the RNA-directed DNA methylation (RdDM) silencing pathway. In maize, loss of mediator of paramutation1 (mop1) encoded RDR2 function results in reactivation of transcriptionally silenced Mu transposons and a substantial reduction in the accumulation of 24 nt short-interfering RNAs (siRNAs) that recruit RNA silencing components.

Laser microdissection-mediated isolation and in vitro transcriptional amplification of plant RNA.

Protocols for laser microdissection and linear amplification of RNA from fixed, sectioned plant tissues are described. When combined with quantitative RT-PCR, microarray analysis, or RNA-sequencing, these procedures enable quantitative analyses of transcript accumulation from microscopic quantities of specific plant organs, tissues, or single cells.

Microdissection of shoot meristem functional domains.

The shoot apical meristem (SAM) maintains a pool of indeterminate cells within the SAM proper, while lateral organs are initiated from the SAM periphery. Laser microdissection-microarray technology was used to compare transcriptional profiles within these SAM domains to identify novel maize genes that function during leaf development. Nine hundred and sixty-two differentially expressed maize genes were detected; control genes known to be upregulated in the initiating leaf (P0/P1) or in the SAM proper verified the precision of the microdissections.

Regulation of small RNA accumulation in the maize shoot apex.

MicroRNAs (miRNAs) and trans-acting siRNAs (ta-siRNAs) are essential to the establishment of adaxial-abaxial (dorsoventral) leaf polarity. Tas3-derived ta-siRNAs define the adaxial side of the leaf by restricting the expression domain of miRNA miR166, which in turn demarcates the abaxial side of leaves by restricting the expression of adaxial determinants. To investigate the regulatory mechanisms that allow for the precise spatiotemporal accumulation of these polarizing small RNAs, we used laser-microdissection coupled to RT-PCR to determine the expression profiles of their precursor transcripts within the maize shoot apex.

Tissue specificity and evolution of meristematic WOX3 function.

The WUSCHEL-related homeobox (WOX) gene PRESSED FLOWER1 (PRS1) performs a conserved function during lateral organ development in Arabidopsis (Arabidopsis thaliana). Expressed in the periphery of the shoot meristem, PRS1 recruits founder cells that form lateral domains of vegetative and floral organs. Null mutations in PRS1 cause the deletion of lateral stipules from leaves and of lateral sepals and stamens from flowers.

Global gene expression analysis of the shoot apical meristem of maize (Zea mays L.).

All above-ground plant organs are derived from shoot apical meristems (SAMs). Global analyses of gene expression were conducted on maize (Zea mays L.) SAMs to identify genes preferentially expressed in the SAM.

T7-based RNA amplification for genotyping from maize shoot apical meristem.

INTRODUCTIONThe use of RNA for genotyping analysis can be advantageous because transcriptomes are significantly smaller than genomes and typically contain far fewer repetitive sequences. Laser capture microdissection (LCM) has been used successfully to isolate sequences (especially rare transcripts) that accumulate in specific tissues. The amount of RNA collected in a standard microdissection is often insufficient for global gene expression analysis but can be increased via linear amplification.

Maize tissue preparation and extraction of RNA from target cells for genotyping.

INTRODUCTIONThe use of RNA for genotyping analysis can be advantageous because transcriptomes are significantly smaller than genomes and typically contain far fewer repetitive sequences. Laser capture microdissection (LCM) has been used successfully to isolate sequences (especially rare transcripts) that accumulate in specific tissues. Where the quantity of isolated material is limiting, amplification can be used to increase the amount of product.

Cell type-specific gene expression profiling in plants by using a combination of laser microdissection and high-throughput technologies.

Laser microdissection (LM) allows for the isolation of specific cells of interest from heterogeneous tissues under direct microscopic visualization with the assistance of a laser beam. By permitting global analyses of gene expression and metabolites in the selected cells, it is a powerful tool for understanding the biological processes in individual cell types during development or in response to various stimuli. Recently, LM technology has been successfully applied to the separation of individual plant cell types.

Diagnostic and prognostic impact of beta-catenin alterations in pediatric liver tumors.

Hepatoblastoma (HBL), a major childhood malignant neoplasm, represents the most frequent malignant liver tumor in childhood. Recent reports have shown the CTNNB1 coding beta-catenin protein to be frequently mutated or deleted at hot-spot regions involving exon 3 in HBL. We investigated the genetic alterations of the CTNNB1 coding beta-catenin protein and expression of several genes downstream of Wnt signals in 4 benign and 17 malignant pediatric liver tumors (PLTs) consisting of 15 HBL, 1 hepatocellular carcinoma, and 1 hepatic immature sarcoma.

Anaconda, a new class of transposon belonging to the Mu superfamily, has diversified by acquiring host genes during rice evolution.

A new type of transposon, named Anaconda (Anac) has been found in rice (Oryza sativa). In this paper, we demonstrate that Anaconda elements have diversified by acquisition of host cellular genes, amplification of the elements, and substitution and deletion of short segments. We identified four Anaconda elements in studies of rice alternative oxidase (AOX) genes, and subsequently isolated an additional 23 elements based on the identity of their terminal inverted repeats (TIRs).

The rice pyruvate decarboxylase 3 gene, which lacks introns, is transcribed in mature pollen.

The rice pyruvate decarboxylase 3 gene (PDC3), which has no introns, was previously postulated to be a pseudogene because no PDC3 mRNA had been detected, even under anaerobic conditions. However, in this study, it was found that rice PDC3 transcripts accumulated in panicles after heading. Within anthers obtained from the panicles, PDC3 was shown to be transcribed in mature pollen by in situ hybridization.

AOX1c, a novel rice gene for alternative oxidase; comparison with rice AOX1a and AOX1b.

A novel gene for alternative oxidase (AOX) was isolated from rice (Oryza sativa L.) and characterized. The deduced amino acid sequence of the novel AOX gene contains features that are conserved among other AOXs.