TRACKING BREEDING SUCCESS OF DEPLOYED TRACKING BREEDING SUCCESS OF DEPLOYED OYSTERS THROUGH MICROSATELLITE VARIATION OYSTERS THROUGH MICROSATELLITE VARIATION

Carlsson J., S.K. Allen Jr, and K.S. Reece

Supported by grants from:NOAA Oyster Restoration Program through the Chesapeake Bay Program Office

The problemRESTORATION EFFORTS HAVE BEEN FOCUSED ON CONSTRUCTING REEFS

SEEDED WITH DOMESTICATED AQUACULTURE STOCKS (MOSTLY DISEASE-RESISTANCE SELCTED STOCKS-DEBY AND CROSBREED LINEs)

ARE THE DEPLOYED DOEMSTICATED OYSTERS CONTRIBUTE SIGNIFICANTLY TO SPAT PRODUCTION?

DEBY DEPLOYMENT – USE mtDNA RFLP (note this also works for Gulf of Mexico strains)

The Question

PRIOR TO DEPLOYMENT

DEPLOYED DEBY

PRODUCED SPAT

  Rare

  BB

  AA

Data from Great Wicomico 2002-2006

CROSBREED DEPLOYMENT – USE?

The Problem 1

Hinf I digest of mtDNA coIII in natural population

Hinf I digest of mtDNA coIII in Crosbreed strain

The coI and coIII mitochondrial genes do not allow for discriminating among non-DEBY or non-Gulf of Mexico strains

The frequency of the BB haplotypes is not 100% in the DEBY line or Gulf strains

An oyster showing the AA haplotype could still be a DEBY or Gulf of Mexico oyster

An oyster showing the BB haplotype has a high probability of being a DEBY or Gulf of Mexico oyster

The Problem 2

DEBY

mtDNA is good for assigning proportion of DEBY/Gulf of Mexico strains but not for assigning individuals to strains

WILD

  Rare

  BB

  AA

The Solution?Microsatellites

+High power

+Highly variable

+High throughput

+Nuclear

-Time consuming to develop

-Expensive Today the most widely used genetic marker for population genetic studies

ATCTATATATATATATATATATATCGTGG

TCGATATATATATATATATATATAGCACC

ATCTATATATATATATATATCGTGG

TCGATATATATATATATATAGCACC

Chromosome from

♀ (TA)10

Chromosome from

♂ (TA)8

Microsatellite

Example 1Example 1

Wild pop1 – blueWild pop2 – whiteDEBY – yellow

Test pop – greyNote!

4 microsatellites4 microsatellites

2 mtDNA genes2 mtDNA genes

Over 85% did Over 85% did significantlysignificantly not assign to DEBY not assign to DEBY

1 individual assigned to DEBY1 individual assigned to DEBY

Factorial Correspondence Analysis (Genetix)

Example 2Example 2

Wild pop 1 – blue

Test pop A – pinkDEBY – yellow

Wild pop 2 - white

Test pop B - grey

4 microsatellites4 microsatellites

2 mtDNA genes2 mtDNA genes

Over 90% did Over 90% did significantlysignificantly not assign to DEBY not assign to DEBY

No individuals assigned to DEBYNo individuals assigned to DEBY

Factorial Correspondence Analysis (Genetix)

Microsatellite Work in ProgressMicrosatellite Work in Progress

Spat-on-shell – deployment of domesticated oysters

Six sites in the Chesapeake Bay

Six samples of wild oysters collected prior to deployment (n=100/site)

Sampling of deployed spat on shell (n=50 per deployment, 3 deployments/site)

Sampling of 8 domesticated aquaculture lines (n=50/line)

Analyze samples with a suite of 8 microsatellite loci

Rappahannock wild – yellow

Deployed spat-on-shell - blue

Example Rappahannock River, Example Rappahannock River, Drumming GroundDrumming Ground

Factorial Correspondence Analysis (Genetix)

Example Rappahannock River, Example Rappahannock River, Drumming GroundDrumming Ground

Hybrids - white

Deployed spat-on-shell - blue

Rappahannock wild – yellow

Factorial Correspondence Analysis of simulated hybrid progeny (Genetix and Hybridlab)

RappannockF1 hybrids Deployed

Hybrid partitioning of simulated hybrid progeny based microsatellite multilocus genotypes (8 loci)

Hybrid partitioning (Structure)

Rappannock F1 hybridsDeployed

Hybrid partitioning (BAPS)

Hybrid partitioning of simulated hybrid progeny based microsatellite multilocus genotypes (8 loci)

Concluding remarksConcluding remarks

-mtDNA RFLP has provided and will continue to provide essential information about the contribution of DEBYs on a group level

-Microsatellites can clearly discriminate between wild and domesticated oysters on a group and on an individual level

-Microsatellites can also be utilized to detect hybrids and to determine the amount of genetic introgression between wild and domesticated oysters (both current and future)

Thanks!Thanks!

Jim WessonJim Wesson

Tommy LeggettTommy Leggett

Wes HeydeckWes Heydeck

LABORATORYLABORATORY

Georgeta ConstantinGeorgeta Constantin

Elizabeth FrancisElizabeth FrancisMissy SouthworthMissy Southworth

P.G. RossP.G. Ross

Mark LuckenbachMark LuckenbachSharon FurinessSharon Furiness

SAMPLINGSAMPLING

Mike CongroveMike Congrove