scholarly journals Laser Capture Microdissection of Uredinia Formed by Melampsora larici-populina Revealed a Transcriptional Switch Between Biotrophy and Sporulation

2010 ◽  
Vol 23 (10) ◽  
pp. 1275-1286 ◽  
Author(s):  
Stéphane Hacquard ◽  
Christine Delaruelle ◽  
Valérie Legué ◽  
Emilie Tisserant ◽  
Annegret Kohler ◽  
...  
Keyword(s):  
Laser Capture Microdissection ◽  
Molecular Mechanisms ◽  
Rust Fungi ◽  
Tissue Preparation ◽  
Cell Defense ◽  
Genome Wide ◽  
Study Gene Expression ◽  
Transcriptional Switch ◽  
Host Infection ◽  
Laser Capture

The foliar rust caused by the basidiomycete Melampsora larici-populina is the main disease affecting poplar plantations in Europe. The biotrophic status of rust fungi is a major limitation to study gene expression of cell or tissue types during host infection. At the uredinial stage, infected poplar leaves contain distinct rust tissues such as haustoria, infection hyphae, and uredinia with sporogenous hyphae and newly formed asexual urediniospores. Laser capture microdissection (LCM) was used to isolate three areas corresponding to uredinia and subjacent zones in the host mesophyll for expression analysis with M. larici-populina whole-genome exon oligoarrays. Optimization of tissue preparation prior to LCM allowed isolation of RNA of good integrity for genome-wide expression profiling. Our results indicate that the poplar rust uredinial stage is marked by distinct genetic programs related to biotrophy in the host palisade mesophyll and to sporulation in the uredinium. A strong induction of transcripts encoding small secreted proteins, likely containing rust effectors, is observed in the mesophyll, suggesting a late maintenance of suppression of host defense in the tissue containing haustoria and infection hyphae. On the other hand, cell cycle and cell defense rescue transcripts are strongly accumulated in the sporulation area. This combined LCM-transcriptomic approach brings new insights on the molecular mechanisms underlying urediniospore formation in rust fungi.

scholarly journals Tissue preparation for laser capture microdissection and RNA extraction from fresh frozen breast tissue

BioTechniques ◽  
2007 ◽  
Vol 43 (1) ◽  
pp. 41-48 ◽  
Author(s):  
Mary Morrogh ◽  
Narciso Olvera ◽  
Faina Bogomolniy ◽  
Patrick I. Borgen ◽  
Tari A. King
Keyword(s):  
Laser Capture Microdissection ◽  
Breast Tissue ◽  
Rna Extraction ◽  
Tissue Preparation ◽  
Fresh Frozen ◽  
Laser Capture

scholarly journals Analysis of Genome-Wide Monoallelic Expression Patterns in Three Major Cell Types of Mouse Visual Cortex Using Laser Capture Microdissection

PLoS ONE ◽  
2016 ◽  
Vol 11 (9) ◽  
pp. e0163663 ◽  
Author(s):  
Chia-Yi Lin ◽  
Shih-Chuan Huang ◽  
Chun-Che Tung ◽  
Chih-Hsuan Chou ◽  
Susan Shur-Fen Gau ◽  
...  
Keyword(s):  
Visual Cortex ◽  
Laser Capture Microdissection ◽  
Expression Patterns ◽  
Cell Types ◽  
Monoallelic Expression ◽  
Genome Wide ◽  
Mouse Visual Cortex ◽  
Laser Capture

scholarly journals Analysis of estrogen-regulated genes in mouse uterus using cDNA microarray and laser capture microdissection

2004 ◽  
Vol 181 (1) ◽  
pp. 157-167 ◽  
Author(s):  
S Ho Hong ◽  
H Young Nah ◽  
J Yoon Lee ◽  
M Chan Gye ◽  
C Hoon Kim ◽  
...  
Keyword(s):  
Cdna Microarray ◽  
Laser Capture Microdissection ◽  
Molecular Mechanisms ◽  
Expression Patterns ◽  
Cell Types ◽  
Integral Membrane Protein ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Mouse Uterus ◽  
Laser Capture

The steroid hormone, estrogen, plays an important role in various physiological events which are mediated via its nuclear estrogen receptors, ERalpha and ERbeta. However, the molecular mechanisms that are regulated by estrogen in the uterus remain largely unknown. To identify genes that are regulated by estrogen, the ovariectomized mouse uterus was exposed to 17beta-estradiol (E2) for 6 h and 12 h, and the data were analyzed by cDNA microarray. The present study confirms previous findings and identifies several genes with expressions not previously known to be influenced by estrogen. These genes include small proline-rich protein 2A, receptor-activity-modifying protein 3, inhibitor of DNA binding-1, eukaryotic translation initiation factor 2, cystatin B, decorin, secreted frizzled-related protein 2, integral membrane protein 2B and chemokine ligand 12. The expression patterns of several selected genes identified by the microarray analysis were confirmed by RT-PCR. In addition, laser capture microdissection (LCM) was conducted to determine the expression of selected genes in specific uterine cell types. Analysis of early and late responsive genes using LCM and cDNA microarray not only suggests direct and indirect effects of E2 on uterine physiological events, but also demonstrates differential regulation of E2 in specific uterine cell types. These results provide a basic background on global gene alterations or genetic pathways in the uterus during the estrous cycle and the implantation period.

Laser microdissection and pressure-catapulting technique to study gene expression in the reoxygenated myocardium

2006 ◽  
Vol 290 (6) ◽  
pp. H2625-H2632 ◽  
Author(s):  
Donald E. Kuhn ◽  
Sashwati Roy ◽  
Jared Radtke ◽  
Sudip Gupta ◽  
Chandan K. Sen
Keyword(s):  
Myocardial Infarction ◽  
Laser Capture Microdissection ◽  
Laser Microdissection ◽  
Pcr Analysis ◽  
Staining Procedure ◽  
Infarct Core ◽  
Spatially Resolved ◽  
Biological Responses ◽  
Study Gene Expression ◽  
Laser Capture

For focal events such as myocardial infarction, it is important to dissect infarction-induced biological responses as a function of space with respect to the infarct core. Laser microdissection pressure catapulting (LMPC) represents a recent variant of laser capture microdissection that enables robot-assisted rapid capture of catapulted tissue without direct user contact. This work represents the maiden effort to apply laser capture microdissection to study spatially resolved biological responses in myocardial infarction. Infarcted areas of the surviving ischemic-reperfused murine heart were identified using a standardized hematoxylin QS staining procedure. Standard staining techniques fail to preserve tissue RNA. Exposure of the tissue to an aqueous medium (typically used during standard immunohistochemical staining), with or without RNase inhibitors, resulted in a rapid degradation of genes, with ∼80% loss in the 1st h. Tissue elements (1 × 104–4 × 106 μm2) captured from infarcted and noninfarcted sites with micrometer-level surgical precision were collected in a chaotropic RNA lysis solution. Isolated RNA was analyzed for quality by microfluidics technology and reverse transcribed to generate high-quality cDNA. Real-time PCR analysis of the cDNA showed marked (200- and 400-fold, respectively) induction of collagen Ia and IIIa at the infarcted site compared with the noninfarcted site. This work reports a sophisticated yet rapid approach to measurement of relative gene expressions from tissue elements captured from spatially resolved microscopic regions in the heart with micrometer-level precision.

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