Project description:Restriction site Associated DNA (RAD) tags are a genome-wide representation of every site of a particular restriction enzyme by short DNA tags. Most organisms segregate large numbers of DNA sequence polymorphisms that disrupt restriction sites, which allow RAD tags to serve as genetic markers spread at a high-density throughout the genome. Here, we demonstrate the applicability of RAD markers for both individual and bulk-segregant genotyping. First, we show that these markers can be identified and typed on pre-existing microarray formats. Second, we present a method that uses RAD marker DNA to rapidly produce a low-cost microarray genotyping resource that can be used to efficiently identify and type thousands of RAD markers. We demonstrate the utility of the former approach by using a tiling path array for the fruit fly to map a recombination breakpoint, and the latter approach by creating and utilizing an enriched RAD marker array for the threespine stickleback. The high number of RAD markers enabled localization of a previously identified region, as well as a second novel region also associated with the lateral plate phenotype. Taken together, our results demonstrate that RAD markers, and the method to develop a RAD marker microarray resource, allow high-throughput, high-resolution genotyping in both model and non-model systems. Keywords: microarray genotyping
Project description:We have analyzed the pattern of gene expression in human primary CD34(+) stem/progenitor cells. We identified 42,399 unique serial analysis of gene expression (SAGE) tags among 106,021 SAGE tags collected from 2.5 x 10(6) CD34(+) cells purified from bone marrow. Of these unique SAGE tags, 21,546 matched known expressed sequences, including 3,687 known genes, and 20,854 were novel without a match. The SAGE tags that matched known sequences tended to be at higher levels, whereas the novel SAGE tags tended to be at lower levels. By using the generation of longer sequences from SAGE tags for gene identification (GLGI) method, we identified the correct gene for 385 of 440 high-copy SAGE tags that matched multiple genes and we generated 198 novel 3' expressed sequence tags from 138 high-copy novel SAGE tags. We observed that many different SAGE tags were derived from the same genes, reflecting the high heterogeneity of the 3' untranslated region in the expressed genes. We compared the quantitative relationship for genes known to be important in hematopoiesis. The qualitative identification and quantitative measure for each known gene, expressed sequence tag, and novel SAGE tag provide a base for studying normal gene expression in hematopoietic stem/progenitor cells and for studying abnormal gene expression in hematopoietic diseases. Keywords: other
Project description:For identifying genes for sex determination in papaya, digital gene expression analysis by Ht-SuperSAGE (Matsumura et al., 2010) was carried out in flowers from male, female and hermaphrodite plants of papaya. Total more than 9,273,744 26bp-tags were obtained by sequence analysis using SOLiD3 and mapped on papaya primitive sex chromosome sequences.
Project description:One of the most recognizable physiological phenomena is the adrenergic-induced fight-or-flight increase in heart rate and cardiac contraction. For the β-adenergic agonist-induced enhancement of calcium influx and transients, and contractility in the heart, we identify the dual requirement of a subpopulation of Rad-bound calcium channels under basal conditions and PKA phosphorylation of Rad. In mice expressing a non-phosphorylatable Rad mutant, basal cardiac contractility is reduced and adrenergic-augmentation of the calcium current and contractility are disabled. Expression of mutant calcium channel β-subunits that cannot bind the mutant Rad restored contractility, revealing a highly specific therapeutic approach to mimic the contractility imparted by adrenergic agonists. Our findings place Rad and its modulation of calcium channels at the nexus of adrenergic modulation of cardiac responses.