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Stickleback fish: Why we have plenty of fish in the sea

HUNTSVILLE, Ala. — New work from the HudsonAlpha Institute for Biotechnology, with collaborators at Stanford University and five other groups, has pinpointed evolution in action.

By determining genomic sequence from many groups of stickleback fish, the scientists were able to show specific genomic changes leading to the ability of different fish populations to adapt to new environments. “We were pleased with the ability of genomics to show us what molecular changes are important in evolutionary processes,” said Richard Myers, Ph.D., president and director of HudsonAlpha.

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At the end of the last ice age, marine stickleback fish were present in many waters, and then became separated into different populations in lakes and streams worldwide. These populations evolved separate traits, such as number of spines, body length or eye size, which allowed them to thrive in their specific habitat.

To tie these traits to specific DNA changes, the scientists generated a reference of the threespine stickleback fish genome at high quality. “With our reference genome and genetic map for stickleback, we will now be able to use it as a model organism for future studies of adaptation and environmental selection,” said HudsonAlpha faculty investigator Jeremy Schmutz.

They then sequenced 21 pairs of fish that varied at different traits and compared them to each other and to the reference fish genome. Small regions of the fish genome stood out due to changes in the genomic DNA, and many of these could be related to how the fish look and behave.

Two interesting findings stood out. First, the changes between fish populations often happened not by mutations in single DNA bases, but by inversions of very large chunks of DNA on fish chromosomes. When these large inversions of DNA occur, fish can no longer breed with each other effectively and start to become separate species.

Second, the scientists saw that when evolution allowing fish to adapt to their environment seemed to come from single DNA base changes, these were most often in regions of the genome that regulate genes and proteins instead of in the genes themselves. In contrast, previous work has shown that in laboratory or domesticated animals, changes in genes and proteins are found more often.

Jane Grimwood, Ph.D., also a faculty investigator at HudsonAlpha, explained, “The predominance of regulatory changes in the evolution of sticklebacks suggests that natural populations may behave differently than domesticated animals, and our genetic mapping of many species will advance similar studies in natural and wild organisms.”

HudsonAlpha researchers have been part of the NHGRI – NIH funded Center for Excellence in Genome Sciences, or CEGS, project for developing stickleback as a model organism since 2002. As part of this effort, they and colleagues at Stanford University have produced genomic resources for several freshwater and marine stickleback fish. Currently, the HudsonAlpha Genome Sequencing Center is building a reference sequence for the Y chromosome for stickleback, as the sequenced fish was a female.

The paper “The genomic basis of adaptive evolution in threespine sticklebacks” was published in the April 4 issue of the journal Nature and is available below.

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Collaborative project between Myers lab and Epicentre featured in Genome Research

December 1, 2011

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A collaborative effort between Epicentre and the HudsonAlpha Institute for Biotechnology resulted in the development of two novel transposon-based methods for RNA-Seq library preparation. The technique, called Tn-RNA-Seq, can use double-stranded cDNA created from rRNA-depleted RNA to prepare an Illumina sequencing library using only two enzymatic reactions. The researchers generated high-quality RNA-Seq libraries from as little as 10 pg of mRNA (~1 ng of total RNA) with this approach.

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They also present a strand-specific RNA-Seq library construction protocol that combines transposon-based library construction with uracil DNA glycosylase and Endonuclease VIII to specifically degrade the second strand constructed during cDNA synthesis. These directional RNA-Seq libraries maintained the same quality as the nondirectional libraries, while showing a high degree of strand specificity (99.5% of reads mapped to the expected genomic strand).

A key benefit of the Tn-RNA-Seq technique is the ability to use extremely low amounts of RNA to prepare high-quality libraries. All six libraries generated using 10 ng to 10 pg of mRNA had at least 72% of aligned reads map to known transcripts, while the library made from 1 pg of mRNA had 62% of aligned reads map to known transcripts. Library complexity was found to be high for all libraries except for the library constructed with 1 pg of mRNA. In general, Tn-RNASeq libraries made with 10 pg or more of mRNA (about 50 cell equivalents) exhibited consistent quality measures. For all libraries except for the library made with 1 pg of mRNA, the rank correlations remained very high (>0.96) indicating highly consistent and reproducible library formation. The directional Tn-RNA-Seq libraries retained the same level of “strandedness” during sequencing compared to libraries made using standard adaptor-ligation methods.

The authors concluded that high-quality RNA-Seq libraries can be constructed efficiently from low input amounts of RNA using the Tn-RNA-Seq methods, and that the procedure is suitable for high-throughput or automated workflows.