Researchers led by bioengineers at the University of California, San Diego have generated the most complete genome sequences from single E. coli cells and individual neurons from the human brain. The breakthrough comes from a new single-cell genome sequencing technique that confines genome amplification to fluid-filled wells with a volume of just 12 nanoliters.
Jeff Gole, a recent bioengineering Ph.D. from University of California, San Diego (photographed) is part of the team that has published a breakthrough single-cell genome sequencing technique that stands to improve our understanding of genomic diversity among cells from the same human brain. With the new approach, the researchers generated the most complete genome sequences published thus far from single E. coli cells and individual neurons from the human brain. The approach, called Microwell Displacement Amplification System, confines genome amplification to fluid-filled wells with a volume of just 12 nanoliters. This work is published in the journal Nature Biotechnology on November 10, 2013. An animated video illustrating the technique is available upon request.
Credit: UC San Diego Jacobs School of Engineering
The study is published in the journal Nature Biotechnology on November 10, 2013.
"Our preliminary data suggest that individual neurons from the same brain have different genetic compositions. This is a relatively new idea, and our approach will enable researchers to look at genomic differences between single cells with much finer detail," said Kun Zhang, a professor in the Department of Bioengineering at the UC San Diego Jacobs School of Engineering and the corresponding author on the paper.
The researchers report that the genome sequences of single cells generated using the new approach exhibited comparatively little "amplification bias," which has been the most significant technological obstacle facing single-cell genome sequencing in the past decade. This bias refers to the fact that the amplification step is uneven, with different regions of a genome being copied different numbers of times. This imbalance complicates many downstream genomic analyses, including assembly of genomes from scratch and identifying DNA content variations among cells from the same individual.
Single-cell Genome SequencingSequencing the genomes of single cells is of great interest to researchers working in many different fields. For example, probing the genetic make-up of individual cells would help researchers identify and understand a wide range of organisms that cannot be easily grown in the lab from the bacteria that live within our digestive tracts and on our skin, to the microscopic organisms that live in ocean water. Single-cell genetic studies are also being used to study cancer cells, stem cells and the human brain, which is made up of cells that increasingly appear to have significant genomic diversity.
For example, the new sequencing approach identified gains or loss of single copy DNA as small as 1 million base pairs, the highest resolution to date for single-cell sequencing approaches. Recent single-cell sequencing studies have used older techniques which can only decipher DNA copy changes that are at least three to six million base pairs.
Amplification in Nano-Scale Wells
The 12 nanoliter (nL) volume microwells in which amplification takes place are some of the smallest volume wells to be used in published protocols for single-cell genome sequencing.
"By reducing amplification reaction volumes 1000-fold to nanoliter levels in thousands of microwells, we increased the effective concentration of the template genome, leading to improved amplification uniformity and reduced DNA contamination," explained Jeff Gole, the first author on the paper. Gole worked on this project as a Ph.D. student in Kun Zhang's bioengineering lab at the UC San Diego Jacobs School of Engineering. Gole is now a Scientist at Good Start Genetics in Cambridge, Mass.
Compared to the most complete previously published single E. coli genome data set, the new approach recovered 50 percent more of the E. coli genome with 3 to 13-fold less sequencing data.
"The results demonstrate that MIDAS provides a much more efficient way to assemble whole bacterial genomes from single cells without culture," the authors write in the Nature Biotechnology paper.
The genomics researchers collaborated with materials science graduate student Yu-Jui (Roger) Chiu on the microfabrication required to create the arrays of microwells. Chiu is working on his Ph.D. in the lab of UC San Diego electrical engineering professor Yu-Hwa Lo, who also directs the Nano3 Labs in UC San Diego's Qualcomm Institute, where microfabrication took place.
"This project would not have succeeded without the fabrication and instrumentation support available at the Jacobs School and the Qualcomm Institute," said Zhang. "We are very excited about our initial results as well as the possibility that researchers around the world will be able to use this approach in many different contexts."
Prof. Kun Zhang is the PI on an NIH-funded center dedicated to the analysis and visualization of RNA transcripts – a proxy for gene activity – from individual cells within the human brain.
This project was funded by US National Institutes of Health grants R01HG004876, R01GM097253, U01MH098977 and P50HG005550, and National Science Foundation grant OCE-1046368.
A patent application has been filed, and UC San Diego is seeking commercial partners to license and develop this innovation into useful products. For information, contact: firstname.lastname@example.org
"Massively parallel polymerase cloning and genome sequencing of single cells using nanoliter microwells," in Nature Technology by: Jeff Gole (1), Athurva Gore (1), Andrew Richards (1), Yu-Jui Chiu (2), Ho-Lim Fung (1), Diane Bushman (3), Hsin-I Chiang (1,5), Jerold Chun (3), Yu-Hwa Lo (4), Kun Zhang (1)
(1) = Department of Bioengineering, Institute for Genomic Medicine and Institute of Engineering in Medicine, University of California, San Diego
(2) = Materials Science and Engineering Program, University of California, San Diego
(3) = Dorris Neuroscience Center, Molecular and Cellular Neuroscience Department, The Scripps Research Institute
(4) = Department of Electrical and Computer Engineering, University of California, San Diego
(5) = Present address: Department of Animal Science, National Chung Hsing University
Daniel Kane | EurekAlert!
Link Discovered between Immune System, Brain Structure and Memory
26.04.2017 | Universität Basel
Researchers develop eco-friendly, 4-in-1 catalyst
25.04.2017 | Brown University
More and more automobile companies are focusing on body parts made of carbon fiber reinforced plastics (CFRP). However, manufacturing and repair costs must be further reduced in order to make CFRP more economical in use. Together with the Volkswagen AG and five other partners in the project HolQueSt 3D, the Laser Zentrum Hannover e.V. (LZH) has developed laser processes for the automatic trimming, drilling and repair of three-dimensional components.
Automated manufacturing processes are the basis for ultimately establishing the series production of CFRP components. In the project HolQueSt 3D, the LZH has...
Reflecting the structure of composites found in nature and the ancient world, researchers at the University of Illinois at Urbana-Champaign have synthesized thin carbon nanotube (CNT) textiles that exhibit both high electrical conductivity and a level of toughness that is about fifty times higher than copper films, currently used in electronics.
"The structural robustness of thin metal films has significant importance for the reliable operation of smart skin and flexible electronics including...
The nearby, giant radio galaxy M87 hosts a supermassive black hole (BH) and is well-known for its bright jet dominating the spectrum over ten orders of magnitude in frequency. Due to its proximity, jet prominence, and the large black hole mass, M87 is the best laboratory for investigating the formation, acceleration, and collimation of relativistic jets. A research team led by Silke Britzen from the Max Planck Institute for Radio Astronomy in Bonn, Germany, has found strong indication for turbulent processes connecting the accretion disk and the jet of that galaxy providing insights into the longstanding problem of the origin of astrophysical jets.
Supermassive black holes form some of the most enigmatic phenomena in astrophysics. Their enormous energy output is supposed to be generated by the...
The probability to find a certain number of photons inside a laser pulse usually corresponds to a classical distribution of independent events, the so-called...
Microprocessors based on atomically thin materials hold the promise of the evolution of traditional processors as well as new applications in the field of flexible electronics. Now, a TU Wien research team led by Thomas Müller has made a breakthrough in this field as part of an ongoing research project.
Two-dimensional materials, or 2D materials for short, are extremely versatile, although – or often more precisely because – they are made up of just one or a...
20.04.2017 | Event News
18.04.2017 | Event News
03.04.2017 | Event News
26.04.2017 | Earth Sciences
26.04.2017 | Health and Medicine
25.04.2017 | Physics and Astronomy