analysis of germination in tomato seeds

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Analysis of Germination in Tomato Seeds Tammy Chan Department of Biology Mentor: Dr. Hiro Nonogaki (Horticulture)

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Analysis of Germination in Tomato Seeds. Tammy Chan Department of Biology Mentor: Dr. Hiro Nonogaki (Horticulture). Importance of tomato seeds. Agriculture and food source. Carries genetic information. Antioxidant. Tomatina. - PowerPoint PPT Presentation

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Page 1: Analysis of Germination in Tomato Seeds

Analysis of Germination in Tomato Seeds

Tammy ChanDepartment of BiologyMentor: Dr. Hiro Nonogaki (Horticulture)

Page 2: Analysis of Germination in Tomato Seeds

Importance of tomato seeds

• Agriculture and food source

•Antioxidant

• Carries genetic information

http://www.bereagardens.org/images/tomato_field.jpgModified from: http://groupenadatoulon.lautre.net/IMG/jpg/Tomato-gene.jpghttp://www.umassvegetable.org/images/soils_crops_pest_mgt/crop/tomato1.jpg

•Tomatina

Modified from: http://upload.wikimedia.org/wikipedia/commons/8/88/Bright_red_tomato_and_cross_section02.jpg

Page 3: Analysis of Germination in Tomato Seeds

Endosperm

Testa

Embryo

Seed Morphology

Page 4: Analysis of Germination in Tomato Seeds

Opposing Forces of Seed Germination

MechanicalResistance

EmbryoGrowth Potential

Page 5: Analysis of Germination in Tomato Seeds

Mechanical Resistance

Testa

Endosperm

- - Man — Man — Man — Man — Man — Man — Man

Gal Gal—

Radicle

Page 6: Analysis of Germination in Tomato Seeds

Endosperm Cap Composition

- - Man — Man — Man — Man — Man — Man — Man

Gal Gal—

β - Mannosidaseβ - Mannanase

α-Galactosidase

Page 7: Analysis of Germination in Tomato Seeds

Sense Antisense

RNAi

Start Stop

DNARNA

Polymerase

Page 8: Analysis of Germination in Tomato Seeds

RNAi

LeMAN2mRNA

antisense

Interference

Hypothesis: The endosperm cap of a tomato seed is weakened by the function of LeMAN2 mannanase during germination.

Page 9: Analysis of Germination in Tomato Seeds

Seed Germination Curve

Seed Germination of Tomato Seeds

0

20

40

60

80

100

120

0 20 40 60 80 100 120

Time (hours)

Seed Germination (%)

Series1

Page 10: Analysis of Germination in Tomato Seeds

Characterization of RNAi plants

• Tissue Printing

To check mRNA:

To check enzyme activity:

•Gel Diffusion Assay

Page 11: Analysis of Germination in Tomato Seeds

Paper towel

Positively-charged membrane

Half seed

Tissue Printing

Page 12: Analysis of Germination in Tomato Seeds

Membrane

RNA

Tissue Printing

AP

Antibody

Detection Substrate

LeMAN2 RNA probe

DIG DIG DIG DIGDIG

Page 13: Analysis of Germination in Tomato Seeds

Tissue Printing

WT Vector RNAi

WT

Vector

RNAi

Page 14: Analysis of Germination in Tomato Seeds

Gel Diffusion Assay

Endosperm Cap

Radicle

Page 15: Analysis of Germination in Tomato Seeds

Gel Diffusion Assay Results

Standard

Wildtype

Vector

RNAi

Page 16: Analysis of Germination in Tomato Seeds

Chen and Bradford (2000) Plant Physiol 124: 1265-1274

Imbibition Time (h)

Pu

nctu

re F

orc

e (

N)

Endosperm cap

Radicle Tip

Texture analyzer

Puncture Force Analysis

WT

RNAi

Page 17: Analysis of Germination in Tomato Seeds

Acknowledgments

Howard Hughes Medical Institute

Dr. Hiro Nonogaki

Dr. Kevin Ahern

Seed Biology Lab

H2 O

Dr. Po-Pu (Eric) LiuDr. Masa AsahinaJessica KristofJing SunJennifer Coppersmith

Jen Hiro JingMe

Page 18: Analysis of Germination in Tomato Seeds