Ancient DNA: Futuristic Technology Brings the Past into the Present

1781140_lIsolating and sequencing DNA from ancient samples is a highly specialized field of research that easily captures the imagination. For me it started in the early 90’s when I read about researchers using PCR (a relatively new technique at the time) to amplify, and subsequently sequence, the mitochondrial DNA of an extinct subspecies of zebra using a sample collected from a skin rug found at an estate in England.

From samples a few hundred years old to ones that are thousands of years old, scientists have made good use of technological advances to push back the boundaries of time. In this video from Science, Evolutionary Biologist Beth Shapiro talks about working with ancient DNA, and how new advances such as Next Generation Sequencing have made it possible to gather more information from ancient samples.

Science put together a Special Issue focused entirely on the research surrounding ancient DNA. You can find all the articles in this Special Issue here:

Special Issue: From mammoths to Neandertals, ancient DNA unlocks the mysteries of the past.

 

Simplifying Next Generation Sequencing Workflow with QuantiFluor® ds-DNA System

DNA SequenceNext-generation sequencing (NGS), also known as high-throughput parallel sequencing, is the all-encompassing term used to describe a number of different modern sequencing technologies. These include Illumina (Solexa) sequencing, Roche 454 sequencing, Ion torrent: Proton / PGM sequencing and SOLiD sequencing to name a few [1].

With the advent of these technologies sequencing DNA and RNA has become much more facile and affordable in comparison to the previously used Sanger sequencing. For these reasons NGS has been the game-changer in the field of modern genomics and molecular biology.

A common starting point for template preparation for NGS platforms is random fragmentation of target DNA and addition of platform-specific adapter sequences to flanking ends. Protocols typically use sonication to shear input DNA, coupled with several rounds of enzymatic modification to produce a sequencer-ready product [2].

Accurate quantification of DNA preparations is essential to ensure high-quality reads and efficient generation of data. Too much DNA can lead to issues such as mixed signals, un-resolvable data and lower number of single reads. Too little DNA, on the other hand, might result in insufficient sequencing coverage, reduced read depth or empty runs, all of which would incur higher costs. The quality of DNA can also vary depending on the source or extraction method applied and further reinforces the need for appropriate management of the input material. Continue reading “Simplifying Next Generation Sequencing Workflow with QuantiFluor® ds-DNA System”

Tips and Tricks for Successful Nucleic Acid Preparation from FFPE Samples: Webinar Preview

FFPE_molecular_analysis_workflowFormalin-fixed, paraffin-embedded (FFPE) tissue samples are extremely common sample types. In this form, tissue is easy to store for extremely long periods of time and useful for immunohistochemical studies. Additionally FFPE samples are fairly inexpensive to produce. However the formalin fixation procedure, which was developed long before the advent of molecular biology, results in chemical crosslinking of nucleic acid and protein molecules inside the cells. This crosslinking presents a challenge for isolating intact, high-quality nucleic acid DNA; so getting at the wealth of molecular information within an FFPE sample can be difficult.

In the upcoming webinar “Successfully Overcoming the Challenges of Working with FFPE Samples”, Dr. Trista Schagat of Promega Corporation discusses some of the key considerations for anyone who is attempting to isolate nucleic acid from FFPE samples. Continue reading “Tips and Tricks for Successful Nucleic Acid Preparation from FFPE Samples: Webinar Preview”

The Power and Potential of Next-Generation Sequencing

DNA in a test tubeNext-generation sequencing (NGS), also known as massively parallel sequencing, is revolutionizing genomic research. NGS technologies have made whole genome sequencing fast and easy, leading to dramatic advances in evolutionary biology and phylogenetics, personalized medicine and forensic science. Why is NGS such a hot topic right now?

Continue reading “The Power and Potential of Next-Generation Sequencing”

DNA Sequencing from AutoRads and Gels to Nanopores

DNA SequencingLast week I read an article in Wired Science that described how an outbreak of antibiotic resistant Klebsiella pneuomiae was tracked in real-time at an NIH hospital using DNA sequencing technologies. The article described how whole genome sequencing of disease isolates and environmental samples from the hospital was used to track the source and spread of the outbreak.

The scientists monitoring the outbreak tracked spontaneous random mutations in the K. pneumoniea DNA sequence to determine that the outbreak was caused by a single source, and to track the spread of the organism within the hospital. The sequencing information helped investigators identify when and where infection occurred, and also to track transmission of the infection from person-to-person. It also revealed that the order of transmission was different from the order in which the cases presented with symptoms, and helped identify how the organism was spread between individuals.

The article describes how epidemiology, infection control and sequence identification were used together to influence outcome in this situation, but also shows the power of whole genome sequencing to find and track subtle differences between isolates that could not have been identified in any other way.

To me, this is a powerful illustration of just how far DNA sequencing has come over the last few years. Not so long ago, the idea of sequencing the entire genome of numerous disease isolates during an outbreak would have been almost laughable—an idea confined to episodes of the X-files or to science fiction stories. Now, thanks to advanced automated sequencing technologies and the computing power to analyze the results, it is doable within a reasonable timeframe for hospitals with access to the right facilities. Although this type of investigation is still beyond the capabilities of most hospitals, the costs and turnaround times for sequencing are coming down rapidly as new technologies capable of faster, cheaper analysis become available.

We have come a very long way since the days when DNA sequencing was a laborious process involving pouring a gel, running samples,and manually reading the resulting autoradiogram hoping to get a read of 50–100 bases. My reading of the wired article prompted me to find out more about the newer types of sequencing technology available today. Here’s what I learned about each: Continue reading “DNA Sequencing from AutoRads and Gels to Nanopores”

Bisulfite Conversion and Next Gen Sequencing

WebinarsIn my last entry, I gave a little summary of one of many techniques that are used to study DNA methylation patterns in a loci-specific fashion using the COBRA technique. This time, we’ll take a look at a high-throughput, genome-wide method for analyzing DNA methylation status using a next generation sequencing approache called bisulfite sequencing, or Bis-Seq. Continue reading “Bisulfite Conversion and Next Gen Sequencing”