DNA sequencing is the process of determining the precise order of nucleotides within a DNA molecule. It includes any method or technology that is used to determine the order of the four bases-adenine, guanine, cytosine, and thymine-in a strand of DNA. The advent of rapid DNA sequencing methods has greatly accelerated biological and medical research and discovery.

Knowledge of DNA sequences has become indispensable for basic biological research, and in numerous applied fields such as medical diagnosis, biotechnology, forensic biology, virology and biological systematics. The rapid speed of sequencing attained with modern DNA sequencing technology has been instrumental in the sequencing of complete DNA sequences, or genomes of numerous types and species of life, including the human genome and other complete DNA sequences of many animal, plant, and microbial species.

There are several DNA sequencing methods and technologies. The chain-termination method developed by Frederick Sanger and coworkers in 1977 became the method of choice for many years, owing to its relative ease and reliability. This was the method used in the first generation of DNA sequencers. Since then, great advances were made in the technique, such as fluorescent labelling, capillary electrophoresis, and general automation. These developments allowed much more efficient sequencing, leading to lower costs. The Sanger method, in mass production form, is the technology which produced the first human genome in 2001, ushering in the age of genomics.

DNA sequencing methods currently under development include reading the sequence as a DNA strand transits through nano-pores, and microscopy-based techniques, such as atomic force microscopy or transmission electron microscopy that are used to identify the positions of individual nucleotides within long DNA fragments. Mass spectrometry may be used to determine DNA sequences. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry, or MALDI-TOF MS, has specifically been investigated as an alternative method to gel electrophoresis for visualizing DNA fragments. With this method, DNA fragments generated by chain-termination sequencing reactions are compared by mass rather than by size. The mass of each nucleotide is different from the others and this difference is detectable by mass spectrometry.

DNA sequencing based on laser-excited fluorescence is characterized by dramatic increases in throughput, reductions in cost, and diverse technological innovations. It is well proven and also the most versatile way of sampling wet chemistry and biochemistry in real time, with unmatched spatial resolution and high (even single-molecule) sensitivity. With advances in chemistry, nano-photonics, and microfluidics, there are many areas where innovation is key within laser technology.

Only a few select wavelengths (405 or 488 nm, 514 nm, 532 nm, and 640 or 660 nm) are required, but diverse choices are needed in every other laser parameter - beam characteristics, output power range, laser core technology (diode/DPSS) etc. For applications where the DNA strands are highly amplified and spatially concentrated, the fluorescence signal is more intense and less laser power is required. But if these sampling sites are then spread over large flow cells or slides needing wide field illumination, that will increase the power requirement. For applications where single strands are being sequenced and signal is therefore inherently low, power is critical. Another factor in the power equation is damage; the laser must not cause functional damage to any of the enzymes used in the sequencing, or to the DNA itself. And because shorter wavelengths are generally more damaging, there is less of a demand for power above a few hundred mw at shorter wavelengths like 405 nm or 488 nm.

Product integration is another key area of innovation for both instrumentation experts and laser technologists. During R&D and product development, at a very minimum instrument developers need smart lasers with plug-and-play simplicity. But they also often need off-the-shelf modular solutions to enable fiber delivery (both single- and multimode), plug-and-play combining of multiple wavelengths, and beam shaping using refractive and diffractive optics. And in some cases, they require an engine in which the coherence of the laser is eliminated in order to avoid speckle and ensure uniform stable illumination. At Laserglow Technologies, we continue our support of this field by offering a wide range of lasers for both proprietary photonics integration and standard illumination requirements.

Sources:

1. "GENETICS/DNA SEQUENCING: Laser fluorescence powers sequencing advances" - Volker Pfeufer and Matthias Schulze BioOptics World

Laserglow's LRS-473 LabSpec Laser was recently used as an illumination source in a research conducted by a team at the Douglas Mental Health University Institute, McGill University, Quebec, Canada to investigate the role of septal glutamatergic neurons in theta rhythm generation in the hippocampus.

The 'hippocampal theta rhythm' is a strong oscillation that can be observed in the hippocampus and other brain structures in numerous species of mammals. It is most notably observed when the subject is engaged in a wakeful-state activity (like running) and during REM sleep. While the precise function of these rhythms is not clearly understood, numerous studies over the decades have indicated that they're closely linked to sensorimotor processing, navigation, learning and memory. This research specifically focused on exploring the functional role of septal glutamatergic neurons, a recently identified population, in theta rhythm generation in the hippocampus. Using optogenetics and electrophysiology, the team explored the functional connectivity of these neurons in vitro, as well as their influence on theta rhythms both in vitro and in vivo, to show that this neuronal population can powerfully drive theta rhythms through intra-septal connections.

The research team believes that their study and observations provide sufficient evidence to suggest that glutamatergic neurons may significantly participate in learning and memory. To learn more about the experimental setup, and for a detailed discussion of the results, go to: Journal of Neuroscience Full Paper

A group of researchers at University of Central Florida has produced the most efficient quantum cascade laser ever designed - and done it in a manner that makes the lasers easier to reproduce. Quantum cascade lasers, or QCLs, are smaller than a grain of rice, and compared to traditional lasers, QCLs offer higher power output and can be tuned to a wide range of infrared wavelengths. They can also be used at room temperature without the need for cooling systems. But due to their size, they're extremely expensive to manufacture.

A University of Central Florida team led by Assistant Professor Arkadiy Lyakh has developed a simpler process for creating such lasers, with comparable performance and better efficiency. The results were published recently in the scientific journal Applied Physics Letters.

"The previous record was achieved using a design that's a little exotic, that's somewhat difficult to reproduce in real life," Lyakh said. "We improved on that record, but what's really important is that we did it in such a way that it's easier to transition this technology to production. From a practical standpoint, it's an important result."

That could lead to greater usage in spectroscopy, such as using the infrared lasers as remote sensors to detect gases and toxins in the atmosphere. Lyakh also envisions QCLs in portable health devices. For instance, a small QCL-embedded device could be plugged into a smartphone and used to diagnose health problems by simply analyzing one's exhaled breath.

The method that previously produced the highest efficiency called for the QCL atop a substrate made up of more than 1,000 layers, each one barely thicker than a single atom. Each layer was composed of one of five different materials, making production challenging. The new method developed at UCF uses only two different materials - a simpler design from a production standpoint. Read more at: Phys.org full article