1: New method for extracting RNA from a single cell without damaging the surrounding tissues
Extraction of RNA from a sample of cells is a routine technique used by molecular biologists around the world. Usually this involves harvesting, and eventually killing cells, breaking them apart before purifying the RNA found within. Some techniques are very sensitive, requiring very little input and they can in fact be used to extract the total RNA of just a single cell. RNA has a variety of applications, primarily in studies of gene expression. Now a study, published in Nature Methods, has demonstrated a technique by which RNA can be extracted from a single cell within intact, live tissue. This technique, which may be the first non-invasive method of single cell RNA extraction, could allow scientists to study the effects of, and a cells response to, changes in the surrounding tissues, known as their microenvironment. There is also good opportunity to study cell-cell communication and chemical connections in much greater detail than previously allowed.
The researchers from the University of Pennsylvania designed and created a molecule, described as a TIVA tag (a transcriptome in vivo analysis tag) that has the ability to enter cells and bind to RNA as it is transcribed. The tags activity is stimulated by a beam of light. Combining this technique with sequencing of the extracted RNA (RNA-seq), the scientists were able to look at the differences in levels of all the RNA in a cell (a cells transcriptome) and compare this to other single cells. This allows for an almost unimaginable level of detail in terms of studying changes in gene expression. Variation in gene expression between cells of the same type is, as the researchers put it, “a well-accepted phenomenon” that has not been studied thoroughly. To make a start on this the scientists here showed that the microenvironment around a neuron “shapes” and modulates the genes expressed.
There are other techniques available that can be used to isolate single cells, and the precious RNA that they contain, from intact tissues, such as combining microdissection and laser capture. But these, and other techniques have a few drawbacks as noted by the researchers in this study. They may involve fixing the tissue (leaving them biochemically inactive, i.e. dead) beforehand, have issues with RNA contamination or even directly cause changes in the make-up of the RNA population. This is generally caused by the intrusiveness of the extraction technique. With the aim of profiling the RNA population of a single cell, any contamination or changes to the RNA would clearly affect the accuracy, reliability and almost all other aspects of the work. This is where the non-invasive properties of the TIVA tag technique come into play and help the method stand out.
There is no defined time course for a new method to be adopted by the scientific community and the ease with which this technique, primarily the production of the TIVA tag, can be replicated and optimised may well be a deciding factor in whether this technique will make the cut.
2: A big week for Illumina and genome sequencing
A suite of HiSeq X tens
Image credit: Illumina
The prospect of the $1000 genome may be within reach (again) according to the CEO of Illumina as he announced a new high-end DNA sequencing platform, the HiSeq X Ten. This $1000 price tag is commonly thought to be the price at which genome sequencing becomes cost-effective for large scale applications in medical research and personalised medicine. It is a price tag that we have heard claimed to be within reach before. Ion Torrent, since acquired by Life Technologies, announced in 2012 that, by 2013, they would have developed a chip that could sequence an entire human genome for $1000. In the case of the HiSeq X Ten the price tag includes the cost of the reaction reagents ($797), the machine itself ($137) and the cost of paying the operators ($55-65) while not including any overhead costs.
Improvements on the previous “high-end” model, HiSeq 2500, offered by Illumina include a better camera, a new set of faster enzymes and improvements to the flow cell (the site of all the reactions) that includes adding nanowells that house an ordered array of specific DNA templates allowing the DNA to be packed into a smaller space (therefore allowing more DNA to be sequenced at once).
Designed for “factory scale” sequencing of (only) human genomes the HiSeq X Ten must be bought as a suite of ten machines coming in at $10m. This could be quite a stretch for many as very few have the need for such high amounts of sequencing. But CEO Jay Flatley pointed out that there were already a number of large scale sequencing projects that could benefit from the increased speed and accuracy offered by the new HiSeq X Ten including the UK’s 100k Genomes project led by Genomics England. The HiSeq X Ten could see itself sequencing tens of thousands of genomes each year, being able to sequence 16 complete genomes in a 3 day run. Although they may be out of reach for many laboratories the first of these machines is set to ship in quarter 1, around March, to customers including Macrogen, the Broad institute and the Garvan Institute of Medical Research in Sydney. Of course Illumina themselves do provide access to whole genome sequencing services through the Illumina Genome Network (IGN) as do a multitude of other organisations who might see a suite of HiSeq X Tens as a good investment.
Another announcement from Illumina this week saw the introduction of their new model of desktop sequencing machine in the NextSeq 500 system. Jay Flatley described “breakthroughs in optics, fluidics and chemistry” which allows increased throughput while still remaining at a “significantly reduced cost” of $250,000. This machine, which can sequence a whole human genome in a day, can sequence both DNA and RNA while also able to be used in the study of microbial genomics.
While both of these offerings from Illumina are an improvement on current models and may help to solidify the company as the primary provider of sequencing technologies, some scientists may still be waiting to see the outcome of other developing technologies such as those of Oxford Nanopore that may provide a novel, faster and cheaper alternative to the current portfolio of next generation sequencers. I highly recommend you check out their website and look at what they plan to offer. It really is some interesting and exciting stuff, with the possibility of an almost USB stick sized sequencer with a variety of other functions as well.
Other good reads this week:
- First find of a black hole orbiting a Be-type “spinning” star
- Chimps shown to use gestures to communicate while hunting for food (Journal article here)
- A bad year for white rhinos in South Africa
- First satellite of the sentinels to launch around early April
DBSci 