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A method for gene expression analysis by oligonucleotide arrays from minute biological materials.

Gene expression profiling has been widely used in identifying differentially expressed genes. One of the most popular formats is oligonucleotide array. A limitation of oligonucleotide arrays is the requirement of relatively large amounts of biological starting materials for gene expression analysis. We have developed a simple method for gene expression profiling from very small amounts of biological material by combining exponential (PCR) and linear (T7 RNA polymerase) amplification. By modifying the widely used SMART protocol, we combined T7 promoter ligation and PCR amplification in one step and generated around 0.5 microg of PCRcDNA from 30 ng of total RNA in a single PCR. The PCRcDNA was in vitro transcribed by T7 RNA polymerase to generate complementary RNA (cRNA), which then was used to hybridize Affymetrix GeneChips. Our results demonstrated a linear correlation between the PCR amplification and the conventional linear amplification in gene expression ratios of individual transcript species between two different RNA preparations. The method was further validated by TaqMan reactions. Moreover, both linear and PCR methods showed some inherent bias as to which transcripts were detected, suggesting that using both in parallel may provide a more comprehensive coverage of the transcriptome present in a given sample.

Base Sequence↗

Integrative genomic and transcriptomic analysis of hypertension in a Taiwanese population.

OBJECTIVES: Hypertension is highly prevalent in Asian populations and represents a major cardiovascular risk factor. However, most genome-wide association studies (GWASs) and transcriptome-wide association studies (TWASs) have focused primarily on Caucasian cohorts. This study aimed to identify genetic loci and gene expression signatures associated with hypertension in an Asian population. METHODS: We analyzed 10 739 hypertensive patients and 49 668 controls from the Taiwan Biobank, testing 4 512 191 genome-wide single nucleotide polymorphisms (SNPs). Integrated GWAS, TWAS, and expression quantitative trait locus (eQTL) analyses were conducted to characterize genetic risk. Additionally, a polygenic risk score (PRS) was constructed using a split-sample design to evaluate genetic risk stratification. RESULTS: We identified 14 loci significantly associated with hypertension, including a novel locus at 5p13.1. eQTL analysis linked this locus to DAB2 expression in whole blood. TWAS detected 55 hypertension-associated genes, with 20 (36%) overlapping GWAS loci. Several novel genes outside GWAS loci, including FBXL15, KCNIP2, and CRIP3, were highly significant and implicated in vascular biology and hypertension mechanisms. PRS analysis effectively differentiated hypertension risk, with individuals in the top 10% showing a > 3.5-fold increased risk compared to the bottom 10%. CONCLUSIONS: Our findings provide new insights into the genetic and transcriptomic landscape of hypertension in Asians. The identification of novel loci and genes advances understanding of disease biology and may guide precision medicine approaches for risk prediction and therapeutic development.

Female↗

Gene expression profiling of cultured human NF1 heterozygous (NF1+/-) melanocytes reveals downregulation of a transcriptional cis-regulatory network mediating activation of the melanocyte-specific dopachrome tautomerase (DCT) gene.

One of the major primary features of the neurocutaneous genetic disorder Neurofibromatosis type 1 are the hyperpigmentary café-au-lait macules where disregulation of melanocyte biology is supposed to play a key etiopathogenic role. To gain better insight into the possible role of the tumor suppressor gene NF1, a transcriptomic microarray analysis was performed on human NF1 heterozygous (NF1+/-) melanocytes of a Neurofibromatosis type 1 patient and NF1 wild type (NF1+/+) melanocytes of a healthy control patient, both cultured from normally pigmented skin and hyperpigmented lesional café-au-lait skin. From the magnitude of gene effects, we found that gene expression was affected most strongly by genotype and less so by lesional type. A total of 137 genes had a significant twofold or more up- (72) or downregulated (65) expression in NF1+/- melanocytes compared with NF1+/+ melanocytes. Melanocytes cultured from hyperpigmented café-au-lait skin showed 37 upregulated genes whereas only 14 were downregulated compared with normal skin melanocytes. In addition, significant genotype xlesional type interactions were observed for 465 genes. Differentially expressed genes were mainly involved in regulating cell proliferation and cell adhesion. A high number of transcription factor genes, among which a specific subset important in melanocyte lineage development, were downregulated in the cis-regulatory network governing the activation of the melanocyte-specific dopachrome tautomerase (DCT) gene. Although the results presented have been obtained with a restricted number of patients (one NF1 patient and one control) and using cDNA microarrays that may limit their interpretation, the data nevertheless addresses for the first time the effect of a heterozygous NF1 gene on the expression of the human melanocyte transcriptome and has generated several interesting candidate genes helpful in elucidating the etiopathology of café-au-lait macules in NF1 patients.

Cells, Cultured↗

Integrated analysis of the genome and the transcriptome by FANTOM.

The key to reliable annotation of a mammalian genome is broad characterisation of the transcriptional output, the transcriptome. FANTOM, the functional annotation of mouse cDNA, is a large-scale analysis of both the genome and the transcriptome of the mouse. In the early days of this work, the transcripts were characterised using our sophisticated methods. After the timely release of the first draft of mouse genome sequences, interesting information was obtained by its integration with these one-by-one annotations. Moreover, each transcript included its expression profile. Here, the two integrated annotation methods used by FANTOM are reviewed: one-by-one and categorised. One-by-one annotation refers to naming carried out based on well-known transcripts or its fragments using the top-down-style pipeline developed mostly by the FANTOM project. Categorised annotation, which refers to transcript grouping, not only helps naming of unknown transcripts, but will be the most utilised method for integration of the genome and the transcriptome from now on.

Abstracting and Indexing↗

Meningioma transcript profiles reveal deregulated Notch signaling pathway.

Meningiomas constitute the second most common central nervous system tumor, and yet relatively little is known about the molecular events that are important for the pathogenesis and malignant progression of these tumors. We have used serial analysis of gene expression to compare the transcriptomes of nonneoplastic meninges and meningiomas of all malignancy grades. A novel finding from this screen is the induction of three components of the Notch signaling pathway: the transcription factor, hairy and enhancer of Split1 (HES1) and two members of the Groucho/transducin-like enhancer of Split family of corepressors, TLE2 and TLE3. TLE corepressors interact and modulate the activity of a wide range of transcriptional regulatory systems, one of which is HES1. We have shown that the transcript and protein levels of HES1, the Notch2 and Notch1 receptors and the Jagged1 ligand are induced in meningiomas of all grades, whereas induction of TLE2 and TLE3 occurs specifically in higher-grade meningiomas. Meningioma cell lines express components of the Notch signaling pathway and an inhibitor of this pathway suppresses meningioma cell survival. These results suggest that deregulated expression of the Notch pathway is a critical event in meningioma pathogenesis and that modulation of this and potentially other signaling pathways by TLE corepressors leads to a more malignant phenotype.

Basic Helix-Loop-Helix Proteins↗

Transcriptome Analysis, Machine Learning, and Experimental Identification of CDK7 Affecting the Progression of Pregnancy-induced Hypertension by Influencing Macrophage Polarization.

INTRODUCTION: Pregnancy-induced hypertension (PIH) is a severe pregnancy complication characterized by placental insufficiency, abnormal vascular remodeling, and immune dysregulation, but personalized therapeutic markers remain unclear. This study aimed to identify key genes and explore immune mechanisms in PIH using transcriptome analysis, machine learning, and experimental validation. METHODS: We analyzed the GSE204835 transcriptomic dataset to screen differentially expressed genes (DEGs) and performed Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome, and Gene Set Enrichment Analysis (GSEA) for functional annotation. Immune infiltration analysis was also performed to examine the immune landscape in PIH. Least Absolute Shrinkage and Selection Operator (LASSO) regression identified key genes, which were validated in a PIH cell model. Flow cytometry and immunofluorescence assays assessed the effect of CDK7 knockdown on macrophage polarization. RESULTS: A total of 1,598 DEGs (1,123 upregulated, 475 downregulated) were identified. Enrichment analyses highlighted associations with embryonic organ development, oxidative phosphorylation, angiogenesis, and oxidative stress. Immune infiltration analysis revealed altered eosinophil and macrophage polarization in PIH. LASSO regression selected 12 key genes, with CDK7 showing the most significant upregulation in the PIH model. CDK7 knockdown promoted macrophage polarization toward the anti-inflammatory M2 phenotype. DISCUSSION: These findings link CDK7 to immune dysregulation in PIH by modulating macrophage polarization, expanding our understanding of PIH's molecular mechanisms. The study's limitations include reliance on public datasets and in vitro models, warranting in vivo validation. CONCLUSION: CDK7 emerges as a potential therapeutic target for PIH, offering new insights into immunoregulatory interventions for this complication.

Female↗

Ancestry and somatic profile predict acral melanoma origin and prognosis.

Acral melanoma, which is not ultraviolet (UV)-associated, is the most common type of melanoma in several low- and middle-income countries including Mexico. Latin American samples are significantly underrepresented in global cancer genomics studies, which directly affects patients in these regions as it is known that cancer risk and incidence may be influenced by ancestry and environmental exposures. To address this, we characterise the genome and transcriptome of 123 acral melanoma tumours from 92 Mexican patients, a population notable because of its genetic admixture. Compared with other studies of melanoma, we found fewer frequent mutations in classical driver genes such as BRAF, NRAS or NF1. While most patients had predominantly Amerindian genetic ancestry, those with higher European ancestry had increased frequency of BRAF mutations and a lower median number of structural variants. The tumours with activating BRAF mutations have a transcriptional profile more similar to cutaneous non-volar melanocytes, suggesting that acral melanomas in these patients may arise from a distinct cell of origin compared to other tumours arising in these locations. KIT mutations were found in a subset of these tumours, and quadruple wild-type samples (non BRAF/NRAS/NF1/KIT) differed from mutated samples in their structural genomic profile and overall and recurrence-free survival patterns. Transcriptional profiling defined three expression clusters; these characteristics were associated with recurrence-free and overall survival. We highlight potential novel low-frequency drivers, such as PTPRJ, NF2 and RDH5. Our study enhances knowledge of this understudied disease and underscores the importance of including samples from diverse ancestries in cancer genomics studies.

Journal Article↗

Derivation and comparative assessment of retinal pigment epithelium from human embryonic stem cells using transcriptomics.

Human stem-cell derivatives are likely to play an important role in the future of regenerative medicine. Evaluation and comparison to their in vivo counterparts is critical for assessment of their therapeutic potential. Transcriptomics was used to compare a new differentiation derivative of human embryonic stem (hES) cells--retinal pigment epithelium (RPE)--to human fetal RPE. Several hES cell lines were differentiated into putative RPE, which expressed RPEspecific molecular markers and was capable of phagocytosis, an important RPE function. Isolated hES cell-derived RPE was able to transdifferentiate into cells of neuronal lineage and redifferentiate into RPE-like cells through multiple passages (>30 Population doublings). Gene expression profiling demonstrated their higher similarity to primary RPE tissue than of existing human RPE cell lines D407 and ARPE-19, which has been shown to attenuate loss of visual function in animals. This is the first report of the isolation and characterization of putative RPE cells from hES cells, as well as the first application of transcriptomics to assess embryonic stem-cell derivatives and their in vivo counterparts--a "differentiomics" outlook. We describe for the first time, a differentiation system that does not require coculture with animal cells or factors, thus allowing the production of zoonoses-free RPE cells suitable for subretinal transplantation in patients with retinal degenerative diseases. With the further development of therapeutic cloning, or the creation of the banks of homozygous human leucocyte antigen (HLA) hES cells using parthenogenesis, RPE lines could be generated to overcome the problem of immune rejection and could be one of the nearest term applications of stem-cell technology.

Cell Differentiation↗

Gene expression profiling of Escherichia coli expressing double Vitreoscilla haemoglobin.

In a recent investigation, expression of a double Vitreoscilla haemoglobin (two fused VHb molecules) in Escherichia coli grown in shake flasks resulted in higher final cell density and considerably higher levels of ribosomes and tRNA. In this study, we have investigated the E. coli transcriptome in cells expressing native VHb, double VHb and control cells lacking VHb by hybridising mRNA from the different constructs to high-density oligonucleotide arrays. Within the 95% confidence interval, 4 and 5% of all detected genes in native VHb cells were up- and down-regulated, respectively; in double VHb cells the corresponding numbers were 6 and 10%, respectively. Dividing the data into different functional groups revealed that genes involved in energy metabolism, central intermediary metabolism and cell processes were the most affected at the mRNA level. Particularly, the up-regulation of genes involved in translation and posttranslational modification observed in double VHb cells demonstrates a strong relationship between the regulation of ribosomal genes and the actual number of ribosomes.

Bacterial Proteins↗

NAD+ Metabolism Licenses Zygotic Genome Activation via PARP7-Mediated ADP-Ribosylation of UHRF1 in Mouse Early Embryos.

Zygotic genome activation (ZGA) is a critical developmental milestone whose metabolic regulation remains unclear. This study identifies a pivotal role for Nicotinamide adenine dinucleotide (NAD+) metabolism in regulating ZGA through poly(ADP‑ribose) polymerase 7(PARP7)-mediated ADP-ribosylation. Using ultra-low input embryo metabolomics, we profiled metabolism from zygote to blastocyst, revealing a significant NAD+ decline at the 2-cell stage. This shift coincided with specific upregulation of the mono-ADP-ribosyltransferase PARP7, confirmed by transcriptomics, quantitative RT-PCR, western blot, and immunofluorescence. Genetic knockdown via trim-away technology or pharmacological inhibition with RBN-2397 caused developmental delay/arrest at the 2-cell stage, impaired blastocyst formation, and defective ZGA. Mechanistically, PARP7 deficiency reduced chromatin accessibility (ATAC-seq), diminished H3K4ac and H3K27ac marks, and impaired RNA polymerase II transcription. Integrated proteomics and ADP-ribosylome analysis of late 2-cell embryos identified UHRF1 as a key PARP7 target, mono-ADP-ribosylated at lysines K30 and K31. This modification stabilized UHRF1 protein (cycloheximide chase), and UHRF1 overexpression partially rescued the transcriptional defects associated with ZGA from PARP7 inhibition. Our findings establish a metabolic-epigenetic axis wherein NAD+ metabolism, via PARP7-mediated ADP-ribosylation of UHRF1, regulates chromatin remodeling and transcriptional activation during ZGA, offering fundamental insights into early development.

Animals↗

Genome-wide identification and comparative analysis of Leucine-Rich Repeat Containing (LRRC) gene and their expression responses to Vibrio alginolyticus infection in the Manila clam (Ruditapes philippinarum).

Leucine-rich repeat (LRR) domains are important components of many pattern recognition receptors (PRRs). Previous studies have demonstrated that LRR domain-containing immune receptors, such as nucleotide-binding oligomerization domain-like receptors (NLRs) and Toll-like receptors (TLRs), play important roles in innate immunity in aquatic animals. In addition to these well-characterized LRR-containing receptors, also possesses a group of LRR-containing proteins. These proteins were collectively referred to as leucine-rich repeat-containing (LRRC) proteins in this study, and their genomic characteristics, evolutionary relationships, were systematically analyzed. In this study, a genome-wide identification and characterization of LRRC genes were performed in the Manila clam. A total of 97 unclassified LRR genes were identified and designated as RpLRRCs.. Expression profiling indicated that RpLRRCs are predominantly expressed in the labial palps, digestive gland, and gills, increasing from the blastula stage and peaking at the juvenile stage during development, based on the transcriptome results from V. alginolyticus, V. anguillarum and V. parahaemolyticus, some RpLRRCs were involved in the response to different Vibrio stress. The qPCR analysis following V. alginolyticus challenge demonstrated that different RpLRRC members exhibit diverse response patterns to Vibrio infection. These results suggest that RpLRRCs may play critical roles in immune regulation. The RpLRRC gene family exhibits diverse structural characteristics and regulatory mechanisms and likely plays important roles in the growth, development, and immune response of R. philippinarum.

Immune response↗

Remodeling of DNA methylation and phenotypic and transcriptional changes in synthetic Arabidopsis allotetraploids.

The joining of different genomes in allotetraploids played a major role in plant evolution, but the molecular implications of this event are poorly understood. In synthetic allotetraploids of Arabidopsis and Cardaminopsis arenosa, we previously demonstrated the occurrence of frequent gene silencing. To explore the involvement of epigenetic phenomena, we investigated the occurrence and effects of DNA methylation changes. Changes in DNA methylation patterns were more frequent in synthetic allotetraploids than in the parents. Treatment with 5-aza-2'-deoxycytidine, an inhibitor of DNA methyltransferase, resulted in the development of altered morphologies in the synthetic allotetraploids, but not in the parents. We profiled mRNAs in control and 5-aza-2'-deoxycytidine-treated parents and allotetraploids by amplified fragment length polymorphism-cDNA. We show that DNA demethylation induced and repressed two different transcriptomes. Our results are consistent with the hypothesis that synthetic allotetraploids have compromised mechanisms of epigenetic gene regulation.

Arabidopsis↗

A hormone-dependent tRNA half promotes cell cycle progression via destabilization of p21 mRNA.

tRNA halves are among the most abundant short non-coding RNAs in the cellular transcriptome. Here we report that in androgen receptor-positive LNCaP prostate cancer cells, the hormone-dependent 5'-tRNALysCUU half promoted cell proliferation by facilitating cell cycle progression. Global mRNA profiling upon the 5'-tRNALysCUU half depletion revealed that the mRNA of p21, a negative regulator of the cell cycle, is post-transcriptionally destabilized via a 5'-tRNALysCUU half-driven mechanism. YBX1, identified as a protein interacting with 5'-tRNALysCUU half in the cytosol, was shown to stabilize p21 mRNA. Specific sequences resembling the 5'-tRNALysCUU half, located in the 3'-UTR of p21 mRNA and termed LL588, were identified as the binding site for YBX1 and are required for p21 mRNA stability. In vitro binding assays demonstrated that the 5'-tRNALysCUU half is capable of displacing YBX1 from LL588. Collectively, our findings suggest that the 5'-tRNALysCUU half directly binds to and displaces YBX1 from p21 mRNA, leading to the destabilization of p21 mRNA and the promotion of cell cycle progression in hormone-dependent cancers. Our study illuminates the role of tRNA halves in regulating mRNA stability and suggests that this may be part of broader regulatory networks affecting mRNA levels, orchestrated by various tRNA halves and their interacting proteins.

Humans↗

Diverse roles for HspR in Campylobacter jejuni revealed by the proteome, transcriptome and phenotypic characterization of an hspR mutant.

Campylobacter jejuni is a leading cause of bacterial gastroenteritis in the developed world. The role of a homologue of the negative transcriptional regulatory protein HspR, which in other organisms participates in the control of the heat-shock response, was investigated. Following inactivation of hspR in C. jejuni, members of the HspR regulon were identified by DNA microarray transcript profiling. In agreement with the predicted role of HspR as a negative regulator of genes involved in the heat-shock response, it was observed that the transcript amounts of 13 genes were increased in the hspR mutant, including the chaperone genes dnaK, grpE and clpB, and a gene encoding the heat-shock regulator HrcA. Proteomic analysis also revealed increased synthesis of the heat-shock proteins DnaK, GrpE, GroEL and GroES in the absence of HspR. The altered expression of chaperones was accompanied by heat sensitivity, as the hspR mutant was unable to form colonies at 44 degrees C. Surprisingly, transcriptome analysis also revealed a group of 17 genes with lower transcript levels in the hspR mutant. Of these, eight were predicted to be involved in the formation of the flagella apparatus, and the decreased expression is likely to be responsible for the reduced motility and ability to autoagglutinate that was observed for hspR mutant cells. Electron micrographs showed that mutant cells were spiral-shaped and carried intact flagella, but were elongated compared to wild-type cells. The inactivation of hspR also reduced the ability of Campylobacter to adhere to and invade human epithelial INT-407 cells in vitro, possibly as a consequence of the reduced motility or lower expression of the flagellar export apparatus in hspR mutant cells. It was concluded that, in C. jejuni, HspR influences the expression of several genes that are likely to have an impact on the ability of the bacterium to successfully survive in food products and subsequently infect the consumer.

Bacterial Proteins↗

Transcriptomic pathology of neocortical microcircuit cell types across psychiatric disorders.

Psychiatric disorders such as major depressive disorder (MDD), bipolar disorder (BD), and schizophrenia (SCZ) are characterized by altered cognition and mood, brain functions that depend on information processing by cortical microcircuits. We hypothesized that psychiatric disorders would display cell type-specific transcriptional alterations in neuronal subpopulations that make up cortical microcircuits: excitatory pyramidal (PYR) neurons and vasoactive intestinal peptide- (VIP), somatostatin- (SST), and parvalbumin- (PVALB) expressing inhibitory interneurons. Using laser capture microdissection followed by RNA sequencing (LCM-seq), we performed cell type-specific molecular profiling of subgenual anterior cingulate cortex, a region implicated in mood and cognitive control. We sequenced libraries from 130 whole cells pooled per neuronal subtype (VIP, SST, PVALB, superficial and deep PYR) in 76 subjects from the University of Pittsburgh Brain Tissue Donation Program, evenly split between MDD, BD and SCZ subjects and healthy controls (totaling 380 bulk transcriptomes from ~50,000 neurons). We identified hundreds of differentially expressed (DE) genes and biological pathways across disorders and neuronal subtypes, with the vast majority in interneurons, particularly PVALB. While DE genes were unique to each cell type, there was a partial overlap across disorders for genes involved in the formation and maintenance of neuronal circuits. We observed coordinated alterations in biological pathways between select pairs of microcircuit cell types, also partially shared across disorders. Finally, DE genes coincided with known risk variants from psychiatric genome-wide association studies, suggesting cell type-specific convergence between genetic and transcriptomic risk for psychiatric disorders. Our study suggests transdiagnostic cortical microcircuit pathology in SCZ, BD, and MDD and sets the stage for larger-scale studies investigating how cell circuit-based changes contribute to shared psychiatric risk.

Humans↗

Integrative TWAS and multi-omics analyses prioritize HSPE1 as a candidate risk gene for bipolar disorder with immune cell-specific regulatory evidence.

BACKGROUND: Bipolar disorder (BD) is a severe psychiatric disorder associated with substantial disability. Although genome-wide association studies have identified multiple BD-associated loci, the underlying genes and mechanisms remain incompletely understood. METHODS: We integrated a European-ancestry BD genome-wide association dataset with cross-tissue and tissue-specific transcriptome-wide association studies (TWAS) and complementary gene-based analysis. Candidate genes were further evaluated using differential expression analysis, consensus clustering, immune infiltration analysis, machine learning, summary-data-based Mendelian randomization, Mendelian randomization using single-cell expression quantitative trait locus data, single-nucleus transcriptomics, phenome-wide association analysis, and virtual screening. RESULTS: The integrative analyses prioritized 37 candidate genes. Peripheral-blood differential-expression analysis identified 14 genes that remained significant after FDR correction, and their expression profiles separated BD samples into two expression-defined clusters. Machine-learning analysis selected UNC50, LMAN2L, LYG2, HSPE1, and KANSL3 for an exploratory classification nomogram. SMR associated genetically predicted higher HSPE1 expression with increased BD risk in two blood eQTL datasets. Cell-type-specific analyses indicated HSPE1-related associations in T-cell and natural killer cell subsets, while single-nucleus analysis descriptively showed higher HSPE1 expression in medial thalamic T cells from BD samples. PheWAS identified no genome-wide significant associations for HSPE1, whereas virtual screening identified candidate compounds with favorable predicted docking scores against the HSPE1 structure. CONCLUSION: This integrative multi-omics study identified HSPE1 as a candidate BD risk gene with immune-cell-related regulatory evidence, providing insight into BD pathogenesis and supporting functional validation.

Humans↗

SAGE profiling and demonstration of differential gene expression along the axial developmental gradient of lignifying xylem in loblolly pine (Pinus taeda).

Wood formation has been studied extensively at the cellular and biochemical levels, but remains poorly understood with respect to gene expression and regulation. As a first step toward identifying genes specifically involved in wood formation and characterizing their roles in determining wood quality, serial analysis of gene expression (SAGE) was used to quantify gene expression in lignifying xylem from a single, 10-year-old loblolly pine (Pinus taeda L.). Two SAGE libraries were generated based on lignifying xylem isolated from either the upper (crown) or lower (base) portions of the trunk. Over 85,000 tags representing a maximum of 27,398 expressed genes were analyzed from the crown wood library, and more than 65,000 tags, representing a maximum of 25,983 expressed genes, were analyzed in the base wood library. Combining these data sets to reflect the sum of genes expressed in lignifying xylem, 150,855 tags were cataloged, representing a maximum of 42,641 different genes. Currently, this study represents the most extensive analysis of its kind in a higher plant and provides a quantitative description of the transcriptome representing the lignifying xylem of a 10-year-old loblolly pine.

DNA, Complementary↗

Microarrays: new tools to unravel parasite transcriptomes.

The ability to monitor the expression levels of thousands of genes in a single microarray experiment is a huge progression from conventional Northern blot analysis or PCR-based techniques. Microarrays can play a pivotal role in the mass screening of genes in a wide range of fields including parasitology. The relatively few parasites that can be readily cultured or isolated from a host, as compared with cell lines or tissue sources, makes microarray technology ideal for maximizing experimental results from a limiting source of starting material. Khan et al. (1999 a) commented in an early review of microarray technology " With this system in place, one can anticipate a time when data from thousands of gene expression experiments will be available for meta-analysis........leading to more robust results and subtle conclusions". Now in 2005, microarrays represent a very powerful resource that can play an important role in the characterization and annotation of the transcriptomes of many parasites of medical and veterinary importance.

Animals↗