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Showing papers by "Arjun Raj published in 2011"


Book ChapterDOI
TL;DR: This protocol describes a method to image individual mRNA molecules in situ using about 50 oligonucleotides complementary to different regions of a target mRNA species using differently colored probe sets for each species.
Abstract: This protocol describes a method to image individual mRNA molecules in situ. About 50 oligonucleotides complementary to different regions of a target mRNA species are used simultaneously. Each probe is labeled with a single fluorescent moiety. When these probes bind to their target, each mRNA molecule becomes so intensely fluorescent that it can be seen as a fine fluorescent spot. Several different mRNA species can be detected in multiplex imaging using differently colored probe sets for each species. An automated image-processing program is used to count the number of mRNA molecules of each species that are expressed in each cell, thus yielding single-cell gene expression profiles.

79 citations


Journal ArticleDOI
TL;DR: It is demonstrated that transgenic expression of DNA methyltransferase 3b in the mouse colon initiates de novo DNA methylation of genes that are similar to genes that become methylated in human colon cancer, consistent with the notion that aberrant methylation in cancer may be attributable to targeting of specific sequences by Dnmt3b rather than to random methylation followed by clonal selection.
Abstract: Human cancer cells frequently have regions of their DNA hypermethylated, which results in transcriptional silencing of affected genes and promotion of tumor formation. However, it is still unknown whether cancer-associated aberrant DNA methylation is targeted to specific genomic regions, whether this methylation also occurs in noncancerous cells, and whether these epigenetic events are maintained in the absence of the initiating cause. Here we have addressed some of these issues by demonstrating that transgenic expression of DNA methyltransferase 3b (Dnmt3b) in the mouse colon initiates de novo DNA methylation of genes that are similar to genes that become methylated in human colon cancer. This is consistent with the notion that aberrant methylation in cancer may be attributable to targeting of specific sequences by Dnmt3b rather than to random methylation followed by clonal selection. We also showed that Dnmt3b-induced aberrant DNA methylation was maintained in regenerating tissue, even in the absence of continuous Dnmt3b expression. This supports the concept that transient stressors can cause permanent epigenetic changes in somatic stem cells and that these accumulate over the lifetime of an organism in analogy to DNA mutations.

66 citations


Journal ArticleDOI
22 Apr 2011-Science
TL;DR: Using carefully constructed experimental systems and sophisticated analyses, they tracked changes in RNA transcription in single live cells and their results provide new details about the dynamics of key steps in transcription.
Abstract: Measurements that involve averaging values from large numbers of cells can obscure the fact that processes such as gene expression are fundamentally dynamic and can vary greatly from cell to cell ( 1 ). Similarly, static snapshots of individual cells can reveal instantaneous differences between cells but cannot reveal often important changes over time; for example, it is impossible to tell whether a yellow light follows a green light using only static pictures of traffic lights. On pages 472 and 475 of this issue, Suter et al. ( 2 ) and Larson et al. ( 3 ) report on using a time-lapse approach to document the dynamics of a key cellular process, RNA transcription. Using carefully constructed experimental systems and sophisticated analyses, they tracked changes in RNA transcription in single live cells. Their results provide new details about the dynamics of key steps in transcription.

3 citations