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Shelly A. Trigg: Hi everyone, my name is Shelley trig and I'm a postdoc in the School of aquatic and fishery Sciences at the University of Washington.

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Shelly A. Trigg: And I'm going to be talking about a culmination of a series of experiments that look at the effects of low pH on the Pacific gooey duck.

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Shelly A. Trigg: Looking at physiology and, you know, mixing DNA methylation, I first want to start by acknowledging the CO authors on these projects that include collaborators from University of Rhode Island. My advisor Stephen Roberts.

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Shelly A. Trigg: Some others from University of Washington and also the Jamestown slalom point Whitney hatchery and tribe.

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Shelly A. Trigg: This work was funded by the foundation for Food and Agriculture Research, which is part of the US Department of Agriculture.

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Shelly A. Trigg: So if you're not familiar with Pacific Glee duck. These are very large burrowing clams and their native to the north eastern Pacific Ocean and span from Alaska down through California

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Shelly A. Trigg: They're pretty important economically and they're one of the most valuable farmed shellfish bringing in over

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Shelly A. Trigg: $20 million in annual sales in Washington State alone. And I just wanted to show these pictures of these are gooey duck seed that are about three millimeters across and they get planted in these

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Shelly A. Trigg: Shores around Puget Sound. And each one of these circles. These are PVC pipes that go down about like foot. So there's one seed planted in each one of those tubes and as the animal grows it digs deeper into the sand until their full size.

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Shelly A. Trigg: And then they're they're sold

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Shelly A. Trigg: And specifically Docker also really important ecologically as they serve as bio filters and a resource for a variety of animals and they're also important for Native American tribes and that they provided sustenance, in the form of food and in revenue.

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Shelly A. Trigg: And the question about how pH effects gooey duck comes from trying to understand how gooey duck might tolerate ocean acidification conditions that are predicted in the future.

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Shelly A. Trigg: As the ocean continues to become lower in pH with climate change.

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Shelly A. Trigg: And there's also questions about if stress conditioning. So, exposing animals to low pH conditions might be beneficial in agriculture. For instance, if

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Shelly A. Trigg: Adults adult animals were exposed to low pH, could they pass on beneficial traits to their offspring, or if juveniles were exposed to environmental stress, could they

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Shelly A. Trigg: Express their resilience as adults.

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Shelly A. Trigg: So to start, uncovering answers to those questions with these this series of experiments looks at rootstock performance and reproductive development.

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Shelly A. Trigg: Juvenile development and also carry over effects or in other words where parents passed down traits to their offspring.

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Shelly A. Trigg: This first experiment looks at how static low pH impacts brood stock. So we had animals reared at an ambient pH and at a low pH or 93 days and I found that the low pH conditioning did affect rootstock survival negatively

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Shelly A. Trigg: And I also found that the low pH condition delayed female reproductive development. So here we're looking at a representative slide from each of the treatment groups and you can see there's a lot more pink.

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Shelly A. Trigg: In the low pH group and that pink is the connective tissue, the white part is follicles. So you can see they're, they're much smaller than in the ambient group and these little purple dots are the eggs. You can see there's much fewer eggs. Then in the ambient group. And now this is this

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Shelly A. Trigg: Data

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Shelly A. Trigg: From all of the these different images.

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Shelly A. Trigg: That we took and this is specifically looking at egg size differences 72 days after the treatment was started, and then

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Shelly A. Trigg: After the treatment had ended, plus an eight day recovery period. So you can see that the low pH group tended to have smaller egg size and the final time point they had pretty similar egg sizes, but interestingly the ambient group.

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Shelly A. Trigg: Showed more

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Shelly A. Trigg: Sort of signs of egg resource option and like the animals had spawned were in the low pH group, we didn't see signs like that. So the animals still continue to develop and we're still deleted development at that later time point now in the males. We didn't see this delay.

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Shelly A. Trigg: They showed very similar proportions of immature sperm and mature sperm and then that continue to develop over time. Similarly,

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Shelly A. Trigg: I'm going to move on and talk about another experiment that was done by Holly Putnam

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Shelly A. Trigg: When she was she's now at University of Rhode Island, and this is work that she did as a postdoc and Stephen Roberts lab.

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Shelly A. Trigg: And what she did is take three month old juveniles and expose them to three different conditions and ambient condition a

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Shelly A. Trigg: Moderate low pH condition and a very low pH condition and she reared them for 23 days. And after that, put them in a common ambient condition for 112 days.

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Shelly A. Trigg: And then split them out into either ambient conditions or low pH conditions and this was to see if any of these prior exposures would impact their response to secondary exposure.

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Shelly A. Trigg: What she found was that after the first exposure animals exposed to the very low pH were smaller in size and the other groups.

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Shelly A. Trigg: Now after the common garden experience she saw that animals exposed to moderate or low pH initially were larger in size. They showed an accelerated growth phenotype.

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Shelly A. Trigg: And then

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Shelly A. Trigg: After the secondary exposure. She saw that animals that had never seen low pH before actually showed a reduced

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Shelly A. Trigg: Rate growth delay they showed a growth delay. So these animals were Ambien animals that never ever saw low pH, even in the secondary exposure.

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Shelly A. Trigg: And these were naive animals that did see low pH and there was a dead they exhibit a growth delay and this is the most interesting thing here is that the moderate and low pH.

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Shelly A. Trigg: Exposed animals did not show

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Shelly A. Trigg: A growth delay upon seeing the secondary exposure. So that indicated there some kind of environmental memory and

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Shelly A. Trigg: To find out the molecular underpinnings of that we sequence the gooey duck genome using 10 X genomics and phase genomics and we found 18 scaffolds spanning 942 mega basis and about 35,000 genes.

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Shelly A. Trigg: And the general methylation landscape.

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Shelly A. Trigg: Looks like this at the blue is methylation. You can see it's densely

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Shelly A. Trigg: Around jeans. So these red blocks or jeans.

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Shelly A. Trigg: I performed a differential methylation analysis.

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Shelly A. Trigg: From animals sampled at different time points and what we're looking at here are just heat maps where the rows each row is a different

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Shelly A. Trigg: differentially math related region, the red indicates high methylation and blue indicates low methylation and on the left side of the heat map is the ambient group, the moderate low pH group and the very low pH group on right

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Shelly A. Trigg: And I'm just showing these to really just show that there are differentially methylation regions and I wanted to know where those regions were the genome. Do they tend to lie and specific features and I found that

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Shelly A. Trigg: The marks that reformed after 10 days of the exposure tend to be in these putative three prime UT are regions and after the secondary exposure. They tend to be in in Toronto and three prime uteri regions.

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Shelly A. Trigg: So now I just want to summarize these results initial low pH exposure leads to increase growth after the common garden and protects against slow growth exhibited by naive clams under re exposure.

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Shelly A. Trigg: DNA methylation occurs primarily in jeans and low pH and do DNA methylation changes tend to be an interest in regions just downstream genes and these methylation changes tend to be in jeans associated with differently.

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Shelly A. Trigg: Lucy's

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Shelly A. Trigg: I'm going to like delete this.

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Shelly A. Trigg: Hi everyone, my name is Shelley trig and I am a postdoc at the University of Washington in the School of aquatic and fisheries sciences.

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Shelly A. Trigg: And I'm going to be talking about a culmination of experiments that look at the impact of low pH on the physiology of Pacific gooey duck.

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Shelly A. Trigg: Before I get started, I want to first acknowledge the CO authors on these experiments which include collaborators at University of Rhode Island. My advisor Stephen Roberts.

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Shelly A. Trigg: Some others from University of Washington and Washington see grant and the Jamestown Scala and point when the hatchery and tribe in this work is funded by the US Department of Agriculture foundation for Food and Agriculture Research

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Shelly A. Trigg: If you're not familiar with the Pacific gooey duck. These are large barring clams that are native

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Shelly A. Trigg: To in the north eastern Pacific Ocean, and they range from Alaska through California and they're among the most valuable farm shellfish on a per acre basis, bringing in over $20 million annually to the state of Washington alone.

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Shelly A. Trigg: This is a picture of gooey duck seed, which are about three millimeters across and that's about the size that they are when they get planted.

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Shelly A. Trigg: Three to five millimeters when they get planted out in these plots. This is like a big flat in Puget Sound. And there are PVC tubes that are about a foot deep. And there is one seed placed in each tube and the tube is to protect it as it grows. And eventually it will

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Shelly A. Trigg: burrow down below the tube and grow into an adult.

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Shelly A. Trigg: Gooey duck are also really important ecologically because they serve as a bio filters and as a prey source for a variety of animals.

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Shelly A. Trigg: And they're important culturally

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Shelly A. Trigg: As they have been for tribes, a source of food and also revenue.

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Shelly A. Trigg: So the question how pH affects gooey duck comes from trying to understand how these animals might fare.

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Shelly A. Trigg: Under ocean acidification conditions that are predicted for the future.

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Shelly A. Trigg: And also if stress conditioning could be beneficial to aquaculture, for instance, if adult animals are exposed to a low pH condition. Could they pass on a resilience to their offspring.

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Shelly A. Trigg: Or if larvae or juveniles are exposed to a low pH condition. Could they be more resilient to low pH as adults.

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Shelly A. Trigg: So to begin, investigating those questions. These experiments that I'm going to talk about look into rootstock performance and reproductive development juvenile development and potential carryover effects or traits that are passed down from parents to offspring.

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Shelly A. Trigg: And this first experiment I looked at the impact of low static low pH on rootstock

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Shelly A. Trigg: And I found that low pH negatively impacted the brute stock survival.

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Shelly A. Trigg: And it also delayed female reproductive development. So here we're looking at images of the gonads from the two different treatment groups and you can see a lot more connective tissue, which is the pink.

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Shelly A. Trigg: A lot fewer eggs, which are the purple dots and smaller follicle sizes which are these white areas.

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Shelly A. Trigg: You can also see at two different time points so

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Shelly A. Trigg: There was a series of

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Shelly A. Trigg: images that were taken after 72 days of treatment. And then at the end point. So you can see there's smaller sizes in the low pH group at the earlier at 72 days after the treatment started

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Shelly A. Trigg: And at the end, there is similar exercise, but in general the ambient groups still looked more developed these animals showed signs of that they had spawned and started resolving the aches.

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Shelly A. Trigg: So the low pH group still develops. They're just delayed and we did not see this effect in the males, the males showed similar proportions of amateurs firm to metric and mature sperm.

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Shelly A. Trigg: At the 72 day time point and continue developing similarly throughout the experiment.

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Shelly A. Trigg: Now I'm going to talk about another experiment that was done by Holly Putnam, who's at University of Rhode Island. Now this is work she did as a postdoc and Stephen Roberts lab.

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Shelly A. Trigg: And what she did was take three month old juvenile clams and expose them to three different pH conditions either ambient moderate low pH or very low pH.

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Shelly A. Trigg: After 23 days under those conditions. She put them all under ambient conditions common garden and after 112 days of those conditions. She split them out into two different groups, either.

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Shelly A. Trigg: ambient exposure or low pH exposure and this was to look at if the initial exposure impacted how they performed after the secondary exposure.

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Shelly A. Trigg: She found that

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Shelly A. Trigg: After the initial exposure animals that were under low pH were smaller in size.

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Shelly A. Trigg: However,

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Shelly A. Trigg: After the common garden, she found that animals that had initially been exposed to low or moderate pH had accelerated growth.

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Shelly A. Trigg: This was almost four months after seeing that exposure. They grew larger in size.

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Shelly A. Trigg: Now, under the secondary low pH exposure animals that had never experienced it before. So, this is this blue dot here is ambient animals that never saw any low pH ever. They were the size. Now Ambien animals that were exposed to.

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Shelly A. Trigg: A low pH during the second period showed a growth delay this growth delay was not seen in animals that had been exposed to low pH in the past.

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Shelly A. Trigg: Indicating that there was some kind of tolerance, the animals gained or some kind of environmental memory, if you will.

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Shelly A. Trigg: And to uncover the molecular mechanisms driving that environmental memory we sequence the gooey duck genome using 10 X genomics and face genomics and found 18 scaffold across 942 mega bases and about 35,000 genes.

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Shelly A. Trigg: We looked at the gooey duck genome methylation and it generally looks like this, where there's high amounts of methylation in within jeans.

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Shelly A. Trigg: I performed a differentially math related region analysis from animals sampled at different time points. And here we're looking at heat maps where each row is a differentially method region.

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Shelly A. Trigg: And the far left column is the ambient group the far right column is the very low pH group in the middle column is the moderate group.

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Shelly A. Trigg: Now, if the areas more read that indicates high methylation and blue indicates low methylation and I'm just showing these to demonstrate that there are

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Shelly A. Trigg: There were differentially mutilated regions that pound, and I wondered where they might be in the genome. If they may lie in particular genome features and I found that the initial exposure tended to have differentially method regions in three prime UTI or were

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Shelly A. Trigg: After the secondary exposure. They also tended to be in three primary Dr or interests.

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Shelly A. Trigg: In general, they were tended to be in within jeans.

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Shelly A. Trigg: And these jeans, happened to be involved in psycho skeleton activity when

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Shelly A. Trigg: We're looking at animals. After the common garden and

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Shelly A. Trigg: An animal's after the secondary exposure, the methylation tended to be in genes involved in nucleic acid binding and mitochondrial activity. So I'm going to move on to a different experiment, looking at the impact of variable low pH on rootstock so here

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Shelly A. Trigg: rootstock are either exposed to ambient or a variable low pH condition.

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Shelly A. Trigg: For 46 days and then they are both put at ambient and you can see that the initial exposure did not affect their survival.

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Shelly A. Trigg: And under ambient conditions they similarly had some had similar survival.

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Shelly A. Trigg: So then, we've looked at the performance of the offspring of these birds. Talk to see if these rootstock may pass on traits.

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Shelly A. Trigg: So we reared their larvae under ambient conditions. This is work that was done by Sam girl who's a grad student in the University of Rhode Island. And here we're looking at size. And you can see over time.

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Shelly A. Trigg: Blue is animals from ambient parents and yellow is animals from variable low pH exposed parents, you can see in general, the size

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Shelly A. Trigg: Is fairly similar.

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Shelly A. Trigg: Hi everyone, my name is Shelley trig and I am a postdoc at the University of Washington in the School of aquatic and fisheries sciences, and I'm going to be talking today about a combination

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Shelly A. Trigg: Of results from a series of experiments looking at

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Shelly A. Trigg: Hi everyone, my name is Shelley trig and I am a postdoc at the University of Washington.

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Shelly A. Trigg: In the school of aquatic and fisheries sciences and today I'm going to be talking about a series of experiments that look at the tolerance of specifically duck to low pH and specifically looking at physiology.

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Shelly A. Trigg: And their genetics and DNA methylation have a fright get started I want to acknowledge the CO authors on these experiments.

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Shelly A. Trigg: Collaborators from University of Rhode Island. My advisor Stephen Roberts some others from University of Washington and Washington see grant and the Jamestown school on point Winnie Patreon tribe.

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Shelly A. Trigg: This work is funded by the US Department of Agriculture foundation for Food and Agriculture Research

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Shelly A. Trigg: If you're not familiar with the Pacific gooey duck. These are large barring plans that are native to the northeastern Pacific and they range from Alaska down through California

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Shelly A. Trigg: And there are among the most valuable farm shellfish on a per acre basis, bringing in over $20 million annually to the state of Washington alone.

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Shelly A. Trigg: I wanted to show this picture of gooey duck seed, which are about three millimeters across these get planted in these PVC tubes that are on these really large plots.

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Shelly A. Trigg: Flat shores of Puget Sound.

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Shelly A. Trigg: And they also serve as

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Shelly A. Trigg: Important ecosystem bio filters and pray sources.

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Shelly A. Trigg: And their gooey duck are also really important culturally as they have sustained tribes by being food and providing revenue.

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Shelly A. Trigg: Hi everyone, my name is Shelley trig and I'm a postdoc from the School of aquatic and fisheries sciences.

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Shelly A. Trigg: Hi everyone, my name is Shelley trig and I'm a postdoc at the University of Washington in the School of aquatic and fisheries sciences.

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Shelly A. Trigg: Today I'm going to be talking about a culmination of a series of experiments that look at the tolerance of Pacific way to low pH and specifically looking at physiology and DNA methylation.

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Shelly A. Trigg: Before I get started, I'd like to acknowledge the CO authors on these experiments which include collaborators from the University of Rhode Island. My advisor Stephen Roberts.

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Shelly A. Trigg: Some others from University of Washington. Washington see grant and the Jamestown slalom point Whitney hatchery and tribe. This work is funded by the US Department of Agriculture foundation for Food and Agriculture Research

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Shelly A. Trigg: If you're not familiar with the Pacific gooey duck. These are large burrowing clams.

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Shelly A. Trigg: That are native to the north eastern Pacific Ocean, and they range from Alaska down through California. There are among the most valuable farm shellfish on a per acre basis and they bring in over $20 million in annual sales.

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Shelly A. Trigg: To Washington State alone. They're about these are going to succeed here and there, about three millimeters across and they get planted in these PVC tubes that are in these shellfish plots.

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Shelly A. Trigg: And they grow down through the tubes into adults.

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Shelly A. Trigg: I need to stop.