Single-cell Whole Genome Analysis (WGA) has recently been accomplished on a variety of high-throughput platforms, including DNA-microarrays, SNP-arrays, and NGS. A key difficulty in the interpretation of single-cell WGA data on any platform is the separation of the numerous WGA artifacts from the genuine genetic variants present in the template genome.
Standard DNA-microarrays can detect copy number variations (CNVs) larger than 2.5 Mb from a single-cell genome [Ref], while targeted array comparative genomic hybridizations can discover approximately 1 Mb-sized DNA imbalances [Ref], although remarkably, CNVs as small as 56 kb in single-cell PCR-based WGA products have been detected [Ref]. Similarly, SNP-arrays can find copy number aberrations encompassing millions of bases in a cell [Ref], but have the advantage of enabling the discovery of copy neutral DNA anomalies and regions of loss-of-heterozygosity (LOH), and allow inferring genome-wide haplotypes [Ref].
Next Generation Sequencing (NGS) has a number of advantages over microarrays enabling improved resolution and accuracy in variant calling [Ref]. First, NGS can examine every nucleotide amplified from the cell and allows genome-wide discovery of the full spectrum of DNA mutations, while microarrays only probe for certain CNV loci.
Secondly, sequencing provides digital precision, with one digital unit representing a mapped sequence read. Finally, paired-end sequencing and mapping discloses the linkage between both ends of each linear DNA-molecule in a sequencing library of a single-cell WGA product, allowing the identification of structural variations via read-pairs mapping discordantly to the reference genome.
Analytical challenges remain in interpreting single-cell NGS data for the full spectrum of genetic variants. Although WGA imperfections due to genome base composition (e.g. %GC-bias) can be computationally corrected for [Ref]. Allelic fractions of heterozygous SNPs [Ref] or aberrantly mapping read pairs following paired-end sequencing of the WGA product [Ref] can be used to increase confidence in CNV measurements. For instance, a real deletion of a diploid locus should show LOH and discordantly mapping read-pairs that explain the DNA loss. Furthermore, the cell cycle stage of the isolated cell must be considered, further complicating the analysis, as cells in S-phase demonstrate a dynamic copy number profile, leading to false structural DNA-imbalance discoveries [Ref].
The identification of the full spectrum of intra- and inter-chromosomal (un)balanced structural variants in a single-cell WGA product is still in its infancy—the main difficulty being to filter true structural variants from chimeric DNA generated during WGA, as well as issues with genome coverage. Although filters have been designed to permit the detection of the structural architecture of DNA copy number variation and even to detect L1-retro-transposition [Ref], many structural variants are still missed in single-cell analyses.
Despite these hurdles, several groups have proven the efficacy of single-cell NGS to detect multiple classes of mutation within a genome and even to detect sister chromatid exchanges following single-cell Strand-seq [Ref]. Step-by-step bioinformatics protocols for analyzing Strand-seq data as well as for copy number profiling single cells through NGS or microarray analysis and commercial solutions (e.g. platforms used within are surfacing.[Ref]
Don't learn science. Play with it Daily News & Analysis By allowing students to undertake experiments and lay their hands on equipment like microscope etc, concepts like magnetism, electricity, osmosis and so on can be made interesting, she says.
The Guardian Nigeria 2013 in review: A year of fluctuating fortunes for science The Guardian Nigeria Microscopic-HIV ACCORDING to a review on science by Nature, from breakthroughs in stem-cell therapies to 'cure' for HIV, the past 12 months have...
Shackleton Crater, a crater on the moon that is a prime target for human exploration may be tantalizingly rich in ice, though researchers warn it could just as well hold none at all.
The scientists investigated Shackleton Crater, which sits almost directly on the moon's south pole. The crater, named after the Antarctic explorer Ernest Shackleton, is more than 12 miles wide (19 kilometers) and 2 miles deep (3 km) — about as deep as Earth's oceans.
The interiors of polar craters on the moon are in nearly perpetual darkness, making them cold traps that researchers have long suspected might be home to vast amounts of frozen water and thus key candidates for human exploration. However, previous orbital and Earth-based observations of lunar craters have yielded conflicting interpretations over whether ice is there.
For instance, the Japanese spacecraft Kaguya saw no discernible signs of ice within Shackleton Crater, but NASA's LCROSS probe analyzed Cabeus Crater near the moon's south pole and found it measured as much as 5 percent water by mass. Now scientists who have mapped Shackleton Crater with unprecedented detail have found evidence of ice inside the crater.
NASA's Lunar Reconnaissance Orbiter essentially illuminated the crater's interior with infrared laser light, measuring how reflective it was. The crater's floor is more reflective than that of other nearby craters, suggesting it had ice.
"Water ice in amounts of up to 20 percent is a viable possibility," study lead author Maria Zuber explained. Don't get your hopes up, though. The amount of ice in Shackleton Crater"can also be much less, conceivably as little as zero," Zuber cautioned.
This uncertainty is due in part to what the researchers saw in the rest of the crater. Bizarrely, while the crater's floor was relatively bright, Zuber and her colleagues observed that its walls were even more reflective. Scientists had thought that if highly reflective ice were anywhere in a crater, it would be on the floor, which live in nearly permanent darkness. In comparison, the walls of Shackleton Crater occasionally see daylight, which should evaporate any ice that accumulates.
These particles are bombarded by solar wind, which knocks the atoms out of order and leaves behind oxygen that is more available to react with hydrogen, for example, to create water molecules. “It is a thrilling possibility ...
Scientists found evidence of wood ash in the center of Qesem Cave in Israel. They removed a chunk of sediment and hardened it in the lab, so they could slice layers and evaluate them under the microscope.
Channel News Asia Geckos in space: Novel robot takes a step to cosmos Channel News Asia A wall-crawling robot inspired by the gecko has taken a small but important step towards a future in space, scientists said on Thursday.
Brain Science National Geographic To tamp down the panic, I breathed smoothly and transported myself to places in my memory, at one point recalling how I had once walked my nine-year-old daughter to school through piles of blizzard snow.
ChicagoNow (blog) Cold weather science projects to make your kids' snow day educational and fun ChicagoNow (blog) I am not a rocket scientist, I was an English major and so this is not hard core science, but these are still interesting science...
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