Explore the vast range of titles available.

Analysis of spatiotemporal patterns of gene expression is an important prerequisite for understanding the molecular basis of embryogenesis. Tissue-specific resolution is desirable, but often not achieved owing to methodical limitations. We used a common model system for embryonic development – the South African clawed frog Xenopus laevis – to demonstrate that laser microdissection and laser-mediated catapulting of tissue samples from histologic sections are feasible even for yolk-rich, fragile embryonic tissue. A combination with RT-PCR provides the possibility of detecting tissue-specific gene expression with high resolution and fidelity. We show that specimens of various sizes and shapes can easily be procured by laser microdissection and pressure catapulting (LMPC). Subsequent RNA-isolation and nested RT-PCR for marker genes revealed that the combination of these methods allows for analysis of specific gene expression in micro-areas. We report on the efficiency and reliability of detection of marker genes in dissected tissue. We further discuss the question of whether such a combination can be applied to certain issues raised in developmental biology with regard to other techniques.

One of the key end points for understanding the molecular basis of embryogenesis is the analysis of spatiotemporal patterns of gene expression. Methodical limitations due to low mRNA levels often prevent a tissue-specific resolution. In this study, we developed an improved laser microdissection technique and RT-PCR that allows marker gene detection in small tissue areas from sections of formalin-fixed paraffin-embedded Xenopus embryos. Tissue pieces were isolated by laser microbeam microdissection and captured by laser pressure catapulting. Neither laser treatment nor conventional histological or immunochemical staining impaired subsequent RNA analysis. Transcripts of tissue-specific marker genes such as endodermin (endoderm), epidermal cytokeratin (epidermal ectoderm), N-CAM (neural tube), myoD (somites), and sonic hedgehog (floor plate) were amplified by nested RT-PCR analysis from small areas of single sections.