Super microscope amplifies Texas A&M researchers look at loblolly pine genetics

Writer: Kay Ledbetter, 806-677-5608, skledbetter@ag.tamu.edu
Contacts: Dr. Nurul Faridi, 979-862-3908, nfaridi@tamu.edu
Dr. Claudio Casola, 979-845-8803, ccasola@tamu.edu
Dr. C. Dana Nelson, 228-832-2747, dananelson@fs.fed.us

COLLEGE STATION – A super microscope may have researchers at the Texas A&M University ecosystem science and management department moving one step closer to writing the final chapter and verse on the genomics of the loblolly pine species.

Dr. Nurul Islam-Faridi, a visiting professor at Texas A&M University, works with the new laser capture microdissection microscope. (Texas A&M AgriLife Communications photo)

Dr. Nurul Faridi, an adjunct professor at Texas A&M University, works with the new laser capture microdissection microscope. (Texas A&M AgriLife Communications photo)

The laser capture microdissection microscope, or LCMM, is among the most versatile and accurate instruments to perform chromosomal work, according to the scientists.

Two scientists at Texas A&M who will utilize the LCMM are Dr. Nurul Faridi, leader of the U.S. Department of Agriculture Forest Service’s Molecular Cytogenetics Laboratory and a collaborating faculty member, and Dr. Claudio Casola, a forest genomics assistant professor with Texas A&M AgriLife Research and the university’s ecosystem science and management department.

Their primarily use of the LCMM, located in the Forest Science Laboratory and the only modern LCMM on the Texas A&M campus, will be to isolate and sequence one chromosome at a time from loblolly pine cells.

The microscope was purchased in a collaborative effort between Faridi’s lab with the U.S. Forest Service’s Southern Institute of Forest Genetics in Saucier, Mississippi, under the direction of Dr. Dana Nelson, and the Texas A&M University System, including AgriLife Research and the ecosystem science and management department.

In addition to supporting basic research, results from the LCMM will also directly benefit the applied loblolly pine breeding effort of the Western Gulf Forest Tree Improvement Program, hosted by the Texas A&M Forest Service and directed by Dr. Thomas Byram.

A microscopic view of a single cell of loblolly pine root tissue shows the 12 chromosome pairs. Chromosomes appear blue after staining with a dye that binds to the DNA. The green and red spots mark specific DNA sequences in different regions of each chromosome pair, helping their identification. (Texas A&M AgriLife Communications photo by Dr. Nurul Islam-Faridi)

A microscopic view of a single cell of loblolly pine root tissue shows the 12 chromosome pairs. Chromosomes appear blue after staining with a dye that binds to the DNA. The green and red spots mark specific DNA sequences in different regions of each chromosome pair, helping their identification. (Texas A&M AgriLife Communications photo by Dr. Nurul Faridi)

Loblolly pine represents the principal southern U.S. commercial forest species, due to its abundance, rapid growth, and pulpwood and lumber value, Casola said. The forest products industry, which mainly relies on loblolly pine, has a large economic impact in Texas – $30.3 billion in industry outputs, supporting more than 130,600 jobs.

Approximately 75 percent of all tree species seedlings planted each year in the U.S. are loblolly pine. The management, conservation and commercial improvement of this pine species are key priorities for forest tree scientists, he said. Understanding the genetic basis of variation and local adaptation in loblolly pine would greatly benefit these activities.

The laser beam of the LCMM, 10 times thinner than a human hair, does not alter the molecular composition of these samples. The scientists said access to this laser beam allows them to collect pure biological samples from tissues, including single cells or chromosomes from any living organism.

“One of the most exciting applications of the LCMM for us is to be able to isolate individual chromosomes from tree and other plant cells,” Faridi said. “Chromosomes are long strings of DNA that carry the genetic information found in every living cell.”

The genome sequence of loblolly pine was recently released, but it is a very large genome that is particularly difficult to assemble, despite having only 12 pairs of chromosomes, he said. Given the size and complexity of this genome, this sequence represents a relatively incomplete draft.

“Proper assembly of the genome will be a big challenge since the genome is extremely large and complex, and contains a very high proportion of repetitive DNA,” Faridi said. “As of now, the current pine genome assembly is highly fragmented. Sequencing individual chromosomes using LCMM would significantly reduce the risk and complexity of the whole genome assembly process.”

Faridi said his Forest Tree Cytogenetics Laboratory has successfully microdissected single and multiple copies of loblolly pine chromosome 12, referred to as Ch 12, which is the smallest and only sub-metacentric chromosome in the genome.

“Eventually we will isolate other chromosomes by tagging fluorescent dyes to specific chromosomes,” he said. “DNA microdissected chromosome samples will be amplified using enzymatic reactions and subsequently sequenced at the Texas A&M Genome Center. We anticipate that these new sequences will enable us to generate high quality assemblies of all loblolly pine chromosomes.”

Casola said this single chromosome assembly process will greatly benefit research by allowing scientists to identify all the estimated 80,000 genes in this species. It also will accelerate the pace of genomic discovery, not only in this species but in all conifer species.

He likened the process to a book: “The initial draft is still missing about one-eighth of the words. We can get the ‘plot,’ if you will, but we cannot understand some paragraphs. Translated back to genomics, this is important because some of these ‘paragraphs’ contain genes, so we are missing some crucial pieces of information.”

Variation in the DNA sequence of loblolly genes ultimately determine the ability of pine trees from different areas to adapt to specific environmental conditions, from temperature to precipitation and soil composition.

“We want to find the exact genetic variants that underlie the trees’ adaptive potential,” Nelson said.

He said identifying the genetic variants can help scientists understand the response of different trees to climate change, develop genetic tests to tell where a tree and its progeny will grow best, and improve knowledge of the genetic basis of important traits in plants. In addition, this work will have similar applications to other crops.

While chromosome capture is the main goal in the loblolly pine genome work, the LCMM applications can range from medicine to agriculture, and include genetic analyses of tissues from tumors and molecular studies of plant pests, Casola said.

“In this regard, the LCMM will be available for the Texas A&M scientific community at large and be a unique instrument on campus,” he said. “We like to think of this microscope as a new powerful tool that will help accelerate discovery and technical advances in a variety of disciplines in our university.”

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