University of Miami and Howard Hughes Medical Institute, Research in Ecology Program, Summer 2008

Saving the Endangered South Florida Slash Pines (Pinus elliottii var. densa)

---- A study of Genetic Variations of Two Slash

Pine Populations

Super Awesome Pines (SAP): Renee Roache, Martin Alexis, Allegra

delValle, Melissa Perez, O’Brian Anderson

University of Miami and Howard Hughes Medical Institute, Research in Ecology Program, Summer 2008

Outline for our talk

• Background on Florida Slash Pine

• Importance of our study

• Methods

• Results

• Discussion

• Acknowledgements

Pinus elliottii var. densa

• Evergreen conifer.• Pollen cones are

lower on the trees than the seed cones.

Pinus elliottii var. densa

• Native to central and south Florida, including the Florida Keys.

• It’s the keystone species in the pine rockland community.

Fire resistant to low

intensity fires

Back Then…

• There used to be 65,000 acres of pine rockland habitat in the 1940’s but now there is less than 1% left.

1940’s Today

Genetic diversity

• Genetic variation is important for populations to adapt to a changing environment

• Such as hurricane

Significant values of South Florida slash pine

Over 225 types of native plants occur Over 225 types of native plants occur here and more than 20% of the plant here and more than 20% of the plant species are found here and nowhere else species are found here and nowhere else in the world. in the world.

Other plants in our study sites

How can we distinguish tree DNA?

Tree 1 Tree 2

AT

CC

GG

GC

CG

GG

CC

AC

TT

C

AG

TG

GC

CC

GG

CC

CG

GA

TA

G

AT

CC

GG

GC

CA

CT

TC

GT

AC

G

TA

CG

AA

GT

GG

CC

CG

GA

TG

C

Marker• We used nuclear

microsatellite markers to study genetic variation.

• Microsatellites are short segments of DNA that have a repeated sequence.

Question:Whether fragmentation has effect

on genetic diversity of South Florida Slash pine?

• Null Hypothesis:

Fragmentation has no effect on genetic diversity of Pinus elliottii var. densa

• Prediction:

Fragmentation decrease genetic diversity.

Study Sites

Everglades National Park

Outside UM campus

Sample Collection10 samples per site

DNA Extraction

DNA Extraction

Polymerase Chain Reaction (PCR)

• PCR is a technique used to amplify a piece of DNA. • PCR enables researchers to produce millions of copies

of a specific DNA sequence in approximately two hours.

- Negative

+ Positive

DNA Gel Electrophoresis

• DNA electrophoresis is an technique used to separate DNA fragments by weight.

• An electric field forces the fragments to migrate from negative to positive.

Gene Scan

• Gene Scan analysis is carried out on a LI-COR 4300 DNA analyzer.

Genotype results at locus PtTX3013 and RPTest01

PtTX3013

RPTest01

S S S

S S S

S: Size Marker, EV: Sample from Everglades, UM: Sample from UM

EV UM

UMEV

EV UM

129/129 -

- 129/129

- -

- 129/129

129/129

129/138 129/129

129/129 -

129/129 129/129

- -

129/129 -

• At both PtTX3013 and RPTest01 locus, there are two different alleles in EV, but only one allele in UM.

• EV and UM share same alleles.• EV have heterozygous trees, all trees are homozygous in

UM, base on the samples we collected.

EV UM

- -

- 126/126

- -

- 126/126

- 126/126

126/126 126/126

123/126 -

126/126 126/126

- -

126/126 -

ResultsGenetic variation of two slash pine populations at locus

PtTX3013 RPTest01

Discussion

• EV and UM share two alleles implies some connectivity between them.

• This may be due to historical connection or possibly dispersal by wind, people or animal.

• Fragmentation increased inbreeding, and decrease heterozygocity.

• Larger population size is more heterozygotic.• Large area population have more genetic

variation than fragmented population.

• Efforts of preservation for such population should be equally applied to all populations, small (UM population) and large (Everglades population).

• Local government should all participate in conservation of the remaining population.

• We cannot afford to sacrifice any populations, even though the population size may be small.

• Small populations still contain alleles of the original population.

Discussion

Acknowledgement We would like to thank Dr. Gaines for organizing

this program, Dr. Krempels for organizing us, Dr. Wang for instructing this research, Mr. Jiang, Ms. Suarez and Mr. Cheso for helping us. We thank our parents and friends, Howard Hughes Medical Institute and University of Miami.