Genotyping for Population Genetic Analysis
Turley, B. et. al. “Multiple drivers of interannual oyster settlement and recruitment in the lower Chesapeake Bay.” Conservation Genetics (2019)
Objective: see if restoration efforts had an effect on the annual recruitment of oysters to the point where genetic contributions could be seen
Utilization of Standard BioTools™ products: • Biomark™ HD
• SNP Type™ panel
• 48.48 Dynamic Array™ IFC for Genotyping
Conclusions: • This study examined whether restoration efforts affected yearly oyster recruitment to uncover genetic contributions
• Our investigation found that one of the two experimental oyster strains recruited new oysters. Two of the NEH hatchery-selected strain's young were found to be hybrids.
• The research showed that the populations from which new oysters come into the river change from year to year in the Lafayette River. It's possible that the flow of water in 2013 was just right for oysters that were born outside of the Lafayette to settle in the
Lafayette, but HABs may have had a bigger effect on post-larval survival in the late summer. In 2014, on the other hand, HABs happened early and weren't as bad.
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OYSTER CASE STUDY
Objective: The objective of this study was to measure gene expression levels of Hemocyte immune response genes when exposed to White Spot Syndrome Virus (WSSV) and Fusarium solani (fungal) in order to understand the molecular mechanisms driving shrimp-pathogen interactions to develop strategies to control disease outbreaks in shrimp production systems with L. vannamei. Utilization of Standard BioTools products: Biomark SystemNanoFlex 4-IFC Controller96.96 dynamic arraysReal-Time PCR Analysis Software Conclusions: Genes that were monitored showed up regulation and down regulation over the time course study. Results reported can serve as immunomarkers for mortality events and could help reduce the loss in shrimp production systems
Methods
• DNA was extracted from sterile gill, mantle and adductor-muscle tissue stored in 95% ethanol
• 95 loci were converted to SNP Type assays using D3 Assay Design software and tested for repeatability
• Oysters were genotyped utilizing 48.48 Dynamic Array IFCs. Before the SNP Type test process, preamplification PCR was done using specific target amplification (STA) primers for each SNP locus to standardize the starting DNA template for each sample.
• The diluted STA products were used as a template on a 48.48 Dynamic Array IFC, which genotypes 47 individual samples plus one control and 48 SNP loci simultaneously
• SNP Genotyping Analysis Software used a k-means clustering approach to handle Biomark HD fluorescence signals
• The genotype of each individual was stable among duplicate runs. However, fluorescence levels varied.
• One of our SNP panel's 48 loci was duplicated to test repeatability
• Before analysis, the duplicate locus was deleted. Reproducibility difficulties limited statistical studies to 41 loci
Background
• Maritime population connectivity is the study of how populations are related and connected
• It is important for the management of species for economic reasons, management of reserve areas and restoration activities • Population connectivity serves as a function of several processes for organisms with a larval planktonic life history strategy, including larval transport, larval dispersal and post-larval survival
• This study examined the connectivity of oyster spat and adult oyster populations, as well as the effects of larval lay, metamorphosis and recruitment. The results of this project can be measured and tested with a variety of methods.
Results
• Gill, mantle and adductor-muscle tissue was collected in a sterile way so that DNA could be taken from it
• Patterns of spat recruitment in the Lafayette River were different from the patterns of where and when post-larval oysters settled, which were based on
shell-string surveys
• All of the shell-string sites were settled at about the same time each year
• Between 2013 and 2014, however, both the time and space patterns were different
• In the 2013 shell-string survey, settlement of
post-larval oysters was first seen in mid-June and steadily rose until early August, when it reached its peak
Conclusions
• Oyster spat from 2013 were genetically more similar to resident adults sampled in the Lafayette River, while the 2014 spat exhibited genotypic frequencies more similar to adults from surrounding rivers. The winds during the spawning seasons differed between years, providing conditions for retention in 2013 and mixing of water masses in 2014.
• We recommend that the monitoring of restoration activities should consider relevant environmental conditions and observe multiple years of recruitment to assess the genetic impacts of restoration plantings and variable reproductive success