This document summarizes research on quantifying genetic variation in two cytological aspects of maize endosperm development: endosperm cell number and the extent of endoreduplication. The researchers studied multiple mapping populations to identify quantitative trait loci (QTL) associated with each trait using flow cytometry and genetic mapping techniques. They identified several QTL for each trait, explaining varying percentages of phenotypic variance. The research provides insights into genetic control of these traits and directions for future studies of endosperm development regulation.
TILLING is a general reverse genetic technique that combines chemical mutagenesis with PCR based screening to identify point mutations in regions of interest.
Genomics, proteomics and metabolomics are the three core omics technologies, which respectively deal with the analysis of genome, proteome and metabolome of cells and tissues of an organism.
TILLING is a general reverse genetic technique that combines chemical mutagenesis with PCR based screening to identify point mutations in regions of interest.
Genomics, proteomics and metabolomics are the three core omics technologies, which respectively deal with the analysis of genome, proteome and metabolome of cells and tissues of an organism.
Targeted Induced Local Lesions IN Genome. Mutations (Single base pair substitution) are created by traditionally used chemical mutagens. Identify SNPs and / or INDELS in a gene / genes of interest from a mutagenized population.
Targeting Induced Local Lesions IN Genomes (TILLING) is a combined tool of plant mutagenesis and DNA Biology to investigate useful mutations at Genomic level. First time used for cotton improvement.
A new era of genomics for plant science research has opened due the complete genome sequencing projects of Arabidopsis thaliana and rice. The sequence information available in public database has highlighted the need to develop genome scale reverse genetic strategies for functional analysis (Till et al., 2003). As most of the phenotypes are obscure, the forward genetics can hardly meet the demand of a high throughput and large-scale survey of gene functions. Targeting Induced Local Lesions in Genome TILLING is a general reverse genetic technique that combines chemical mutagenesis with PCR based screening to identity point mutations in regions of interest (McCallum et al., 2000). This strategy works with a mismatch-specific endonuclease to detect induced or natural DNA polymorphisms in genes of interest. A newly developed general reverse genetic strategy helps to locate an allelic series of induced point mutations in genes of interest. It allows the rapid and inexpensive detection of induced point mutations in populations of physically or chemically mutagenized individuals. To create an induced population with the use of physical/chemical mutagens is the first prerequisite for TILLING approach. Most of the plant species are compatible with this technique due to their self-fertilized nature and the seeds produced by these plants can be stored for long periods of time (Borevitz et al., 2003). The seeds are treated with mutagens and raised to harvest M1 plants, which are consequently, self-fertilized to raise the M2 population. DNA extracted from M2 plants is used in mutational screening (Colbert et al., 2001). To avoid mixing of the same mutation only one M2 plant from each M1 is used for DNA extraction (Till et al., 2007). The M3 seeds produce by selfing the M2 progeny can be well preserved for long term storage. Ethyl methane sulfonate (EMS) has been extensively used as a chemical mutagen in TILLING studies in plants to generate mutant populations, although other mutagens can be effective. EMS produces transitional mutations (G/C, A/T) by alkylating G residues which pairs with T instead of the conservative base pairing with C (Nagy et al., 2003). It is a constructive approach for users to attempt a range of chemical mutagens to assess the lethality and sterility on germinal tissue before creating large mutant populations.
Nitrogen Use Efficiency (NUE ) is defined as the yield obtained per unit of available nitrogen (N) in the soil
Efficiency with which the plant uses N from acquired available N to total plant dry matter
NUE is the product of uptake efficiency and utilization efficiency
It is required in all environmental conditions where ever yield of plant is required , NUE -> crop yield
To minimize N loss, maximize N uptake and reduce environmental pollution
NUE is a complex quantitative traits which involves many genes
Expression of multiple gene depend on a number of internal and external factors
There are 100s of nitrate responsive genes
For their transcription require regulatory sequence i.e., NRE (Nitrate responsive elements)
One such element originally reported to be comprised of an A[G/C]TCA sequence
These sequence are randomly distributed throughout the genome
QTL mapping is a powerful tool for analysis of complex NUE
Candidate genes encoding enzyme that involved in N uptake, assimilation and utilization have been reported in rice, maize, arabidopsis etc
Genetic variation and phenotypic plasticity for NUE
Determine the level of genetic variation - landraces and genotypes
Study a defined genetic population under different N conditions
Interactions between N uptake and water availability
Interaction between different macronutrients and micronutrients
Genotype by environment (G × E) interaction
Modifying the root system
Next generation genomics for chickpea (Cicer arietinum L.) improvementICRISAT
Large scale genomic resources including draft genome sequence, re-sequencing of 90 lines, comprehensive transcriptome assembly and high density genetic maps have been developed for chickpea. Linkage mapping and genome wide association studies (GWAS) are being used for trait
mapping.
26 Feb 2014
Extranuclear inheritance or cytoplasmic inheritance is the transmission of genes that occur outside the nucleus. It is found in most eukaryotes and is commonly known to occur in cytoplasmic organelles such as mitochondria and chloroplasts or from cellular parasites like viruses or bacteria. Determining the contribution of organelle genes to plant phenotype is hampered by several factors, including the paucity of variation in the plastid and mitochondrial genomes. Mitochondria are organelles which function to transform energy as a result of cellular respiration. Chloroplasts are organelles which function to produce sugars via photosynthesis in plants and algae. The genes located in mitochondria and chloroplasts are very important for proper cellular function, yet the genomes replicate independently of the DNA located in the nucleus, which is typically arranged in chromosomes that only replicate one time preceding cellular division. The extranuclear genomes of mitochondria and chloroplasts however replicate independently of cell division. They replicate in response to a cell's increasing energy needs which adjust during that cell's lifespan. There is consistent difference between the results from reciprocal crosses; generally only the trait from female parent is transmitted. In most cases, there is no segregation in the F2 and subsequent generations.
Plant genetic engineering is one of the key technologies for crop improvement as well as an emerging approach for producing recombinant proteins in plants. Both plant nuclear and plastid genomes can be genetically modified, yet fundamental functional differences between the eukaryotic genome of the plant cell nucleus and the prokaryotic-like genome of the plastid will have an impact on key characteristics of the resulting transgenic organism. So, which genome, nuclear or plastid, to transform for the desired transgenic phenotype? In this paper we compare the advantages and drawbacks of engineering plant nuclear and plastid genomes to generate transgenic plants with the traits of interest, and evaluate the pros and cons of their use for different biotechnology and basic research applications. The chloroplast is a pivotal organelle in plant cells and eukaryotic algae to carry out photosynthesis, which provides the primary source of the world’s food. The expression of foreign genes in chloroplasts offers several advantages over their expression in the nucleus: high-level expression, no position effects, no vector sequences allowing stable transgene expression. In addition, transgenic chloroplasts are generally not transmitted through pollen grains because of the cytoplasmic localization. In the past two decades, great progress in chloroplast engineering has been made.
Genome projects and their ContributionsAlbertPaul18
This is a presentation about different Genome projects like Rice genome project, Maize genome project, Wheat Genome project and Human genome project. It highlights how they were conducted and what the science community gained by conducting them. A side about the future challenges of such genome projects is also added.
Targeted Induced Local Lesions IN Genome. Mutations (Single base pair substitution) are created by traditionally used chemical mutagens. Identify SNPs and / or INDELS in a gene / genes of interest from a mutagenized population.
Targeting Induced Local Lesions IN Genomes (TILLING) is a combined tool of plant mutagenesis and DNA Biology to investigate useful mutations at Genomic level. First time used for cotton improvement.
A new era of genomics for plant science research has opened due the complete genome sequencing projects of Arabidopsis thaliana and rice. The sequence information available in public database has highlighted the need to develop genome scale reverse genetic strategies for functional analysis (Till et al., 2003). As most of the phenotypes are obscure, the forward genetics can hardly meet the demand of a high throughput and large-scale survey of gene functions. Targeting Induced Local Lesions in Genome TILLING is a general reverse genetic technique that combines chemical mutagenesis with PCR based screening to identity point mutations in regions of interest (McCallum et al., 2000). This strategy works with a mismatch-specific endonuclease to detect induced or natural DNA polymorphisms in genes of interest. A newly developed general reverse genetic strategy helps to locate an allelic series of induced point mutations in genes of interest. It allows the rapid and inexpensive detection of induced point mutations in populations of physically or chemically mutagenized individuals. To create an induced population with the use of physical/chemical mutagens is the first prerequisite for TILLING approach. Most of the plant species are compatible with this technique due to their self-fertilized nature and the seeds produced by these plants can be stored for long periods of time (Borevitz et al., 2003). The seeds are treated with mutagens and raised to harvest M1 plants, which are consequently, self-fertilized to raise the M2 population. DNA extracted from M2 plants is used in mutational screening (Colbert et al., 2001). To avoid mixing of the same mutation only one M2 plant from each M1 is used for DNA extraction (Till et al., 2007). The M3 seeds produce by selfing the M2 progeny can be well preserved for long term storage. Ethyl methane sulfonate (EMS) has been extensively used as a chemical mutagen in TILLING studies in plants to generate mutant populations, although other mutagens can be effective. EMS produces transitional mutations (G/C, A/T) by alkylating G residues which pairs with T instead of the conservative base pairing with C (Nagy et al., 2003). It is a constructive approach for users to attempt a range of chemical mutagens to assess the lethality and sterility on germinal tissue before creating large mutant populations.
Nitrogen Use Efficiency (NUE ) is defined as the yield obtained per unit of available nitrogen (N) in the soil
Efficiency with which the plant uses N from acquired available N to total plant dry matter
NUE is the product of uptake efficiency and utilization efficiency
It is required in all environmental conditions where ever yield of plant is required , NUE -> crop yield
To minimize N loss, maximize N uptake and reduce environmental pollution
NUE is a complex quantitative traits which involves many genes
Expression of multiple gene depend on a number of internal and external factors
There are 100s of nitrate responsive genes
For their transcription require regulatory sequence i.e., NRE (Nitrate responsive elements)
One such element originally reported to be comprised of an A[G/C]TCA sequence
These sequence are randomly distributed throughout the genome
QTL mapping is a powerful tool for analysis of complex NUE
Candidate genes encoding enzyme that involved in N uptake, assimilation and utilization have been reported in rice, maize, arabidopsis etc
Genetic variation and phenotypic plasticity for NUE
Determine the level of genetic variation - landraces and genotypes
Study a defined genetic population under different N conditions
Interactions between N uptake and water availability
Interaction between different macronutrients and micronutrients
Genotype by environment (G × E) interaction
Modifying the root system
Next generation genomics for chickpea (Cicer arietinum L.) improvementICRISAT
Large scale genomic resources including draft genome sequence, re-sequencing of 90 lines, comprehensive transcriptome assembly and high density genetic maps have been developed for chickpea. Linkage mapping and genome wide association studies (GWAS) are being used for trait
mapping.
26 Feb 2014
Extranuclear inheritance or cytoplasmic inheritance is the transmission of genes that occur outside the nucleus. It is found in most eukaryotes and is commonly known to occur in cytoplasmic organelles such as mitochondria and chloroplasts or from cellular parasites like viruses or bacteria. Determining the contribution of organelle genes to plant phenotype is hampered by several factors, including the paucity of variation in the plastid and mitochondrial genomes. Mitochondria are organelles which function to transform energy as a result of cellular respiration. Chloroplasts are organelles which function to produce sugars via photosynthesis in plants and algae. The genes located in mitochondria and chloroplasts are very important for proper cellular function, yet the genomes replicate independently of the DNA located in the nucleus, which is typically arranged in chromosomes that only replicate one time preceding cellular division. The extranuclear genomes of mitochondria and chloroplasts however replicate independently of cell division. They replicate in response to a cell's increasing energy needs which adjust during that cell's lifespan. There is consistent difference between the results from reciprocal crosses; generally only the trait from female parent is transmitted. In most cases, there is no segregation in the F2 and subsequent generations.
Plant genetic engineering is one of the key technologies for crop improvement as well as an emerging approach for producing recombinant proteins in plants. Both plant nuclear and plastid genomes can be genetically modified, yet fundamental functional differences between the eukaryotic genome of the plant cell nucleus and the prokaryotic-like genome of the plastid will have an impact on key characteristics of the resulting transgenic organism. So, which genome, nuclear or plastid, to transform for the desired transgenic phenotype? In this paper we compare the advantages and drawbacks of engineering plant nuclear and plastid genomes to generate transgenic plants with the traits of interest, and evaluate the pros and cons of their use for different biotechnology and basic research applications. The chloroplast is a pivotal organelle in plant cells and eukaryotic algae to carry out photosynthesis, which provides the primary source of the world’s food. The expression of foreign genes in chloroplasts offers several advantages over their expression in the nucleus: high-level expression, no position effects, no vector sequences allowing stable transgene expression. In addition, transgenic chloroplasts are generally not transmitted through pollen grains because of the cytoplasmic localization. In the past two decades, great progress in chloroplast engineering has been made.
Genome projects and their ContributionsAlbertPaul18
This is a presentation about different Genome projects like Rice genome project, Maize genome project, Wheat Genome project and Human genome project. It highlights how they were conducted and what the science community gained by conducting them. A side about the future challenges of such genome projects is also added.
Presentation delivered by Dr. Jesse Poland (Kansas State University, USA) at Borlaug Summit on Wheat for Food Security. March 25 - 28, 2014, Ciudad Obregon, Mexico.
http://www.borlaug100.org
When breeding diploid potatoes, tetraploid progeny can result from the union of 2n eggs and 2n pollen in 2x-2x crosses. Thirty-three crosses were made to examine tetraploid progeny frequency in 2x-2x crosses. All crosses were between S. tuberosum dihaploids and diploid self-compatible donors, M6 and DRH S6-10-4P17. Using chloroplast counting for ploidy determination, the frequency of tetraploid progeny was as high as 45% in one of the 33 crosses. Based upon single nucleotide polymorphism (SNP) genotyping, the tetraploid progeny were attributed to bilateral sexual polyploidization (BSP), which is caused by the union of 2n egg and 2n pollen. Dihaploids were identified that produce lower frequencies of 2n eggs. The results of this study suggest that S. tuberosum dihaploids with a high frequency of 2n eggs should be avoided in 2x - 2x crosses for diploid breeding programs.
'Genomics' is nothing but the study of entire genetic compliment of an organism. Plant genomics is study of plant genome. This is my topic of M.Sc. course 'Plant biotechnology'.
Genome project of Human and methods of sequencing human genome; Genome project of Rice and its post genome sequencing era; Arabidopsis genome project: Why Rice and Arabidopsis chosen for genome project?
The number of sequenced genes having unknown function continues to climb with the continuing decrease in the cost of genome sequencing. In Reverse Genetics (RG), functions of known genes are investigated with targeted modulation of gene activity, and hypothesis regarding gene function directly tested in vivo. Several RG approaches like insertional mutagenesis, fast neutron mutagenesis, TILLING and RNA interference have led to the identification of mutations in candidate genes and subsequent phenotypic analysis of these mutants.
Okabe et al. (2011) employed TILLING technique to screen six ethylene receptor genes in tomato (SlETR1–SlETR6) and two allelic mutants of SlETR1 (Sletr1-1 and Sletr1-2) with reduced ethylene response were identified. Using fast neutron mutagenesis, Li et al. (2001) obtained arabidopsis deletion mutants for bZIP transcription factor viz. AHBP 1b and OBF 5, a key regulator for systemic acquired resistance but their role were compensated by other regulatory factors in mutants. Terada et al. (2007) successfully blocked the expression of the Adh 2 gene through homologous recombination followed by transgenesis in rice however phenotype could not be determined since no differences were observed between wild and transgenic plants. RNA interference (RNAi) works as sequence-specific gene regulation and has been used in determination of function of many genes. Saurabh et al. (2014) reviewed the impact of RNAi in crop improvement and found its application in improvement of nutritional aspects, biotic and abiotic stresses, morphol¬ogy, crafting male sterility, enhanced secondary metabolite synthesis.
In addition, new advances in technology and reduction in sequencing cost may soon make it practical to use whole genome sequencing or gene targeting like ZFN technology and TAL effectors technology on a routine basis to identify or generate mutations in specific genes. Scholze and Boch (2011) mentioned that TAL effectors technology is more specific and predictable than ZFN. RG techniques have their own advantages and disadvantages depending on the species being targeted and the questions being addressed. Finally, with the continuous development of new technologies, the most efficient RG technique in the future may involve high throughput direct sequencing of part or complete genomes of individual plants followed by efficient novel tools to determine the function for utilization in crop improvement.
1. Genetic Analysis of Variation
in Endosperm Cell Number
and
Endoreduplication in Maize
(Zea Mays L.)
2. Outline
•Introduction and goal of research
•Study of two cytological aspects of endosperm development
• Endosperm cell number (mitosis control)
• Extent of endoreduplication (alternative cell cycle
control)
•Materials and methods
• Genetic mapping populations and techniques
• Flow cytometry
•Results
• Heritability estimates
• Genetic correlation estimates
• QTL localization and estimation of genetic effect
• Endosperm development of Zea diploperennis
•Challenges and thoughts for future research
•Conclusions
3. Why Study the Cytological
Aspects of Endosperm
Development?
• Endosperm composes 80-85% of the mature kernel
• ~ 65% Starch and 15% protein (dry weight basis)
• Control of endosperm cell growth
•Mitosis (cell number)
•Endoreduplication (nuclei DNA content)
•Plant improvement
•Quantitative genetics of endosperm growth
•Discover novel genes to increase:
•Grain quality
•Grain yield
4.
5. Biological Significance of the Endoreduplication Cycle
•Common in tissues with high metabolic activity and is often
associated with high levels of gene expression.
•Possibly functions to supply sufficient template for
rapid development.
•May function to provide phosphate and nucleotides for
embryo development and/or the germinating kernel.
•Skeletal function of non-genic nuclear DNA:
Increased DNA required for balanced growth. Serves to
maintain the overall cyto-nuclear ratio in cells that
have increased in volume.
(T. Cavalier-Smith and M.J. Beaton, Genetica 106:
3-13, 1999)
•Faster, more efficient development (Dead End Tissue)
Mitosis (RNA transcription is halted), endocycle has deleted
the mitotic phase.
7. Endosperm Development
Starchy Maize
0 30 6050402010
S C D E Reserve Deposition Dry
Down,
Mature
D.A. DeMason 1997
Cellular and Molecular Biology
of Plant Seed Development
S- Syncytial Phase
C- Cellularization
D- Differentiation and Mitosis
E- Endoreduplication
Days after pollination
Note overlap of events, not
precise demarcations.
8. Larkins et al. 2001
J. Exp. Bot. Vol. 52
Stain:
Propidium
iodide
Growth Rate:
Dry matter
accumulation
from 1 mg
kernel−1 · day−1
(at pollination)
to
6 mg
kernel−1 · day−1
(post 15 DAP)
(Ingle et al.,
1965;
Zinselmeier et
al., 1999.)
11. •N: haploid
Chr. set
•x: chromatid
number per
chromosome
•C: nuclear
DNA
content
N multiplied
By x gives C
12. Materials And Methods
•Two recombinant inbred line (RIL) mapping populations
(Lines previously developed by taking an F1 line through
multiple rounds of selfing: 11+ generations;
essentially a fixed genotype with little or no
within-line genetic variance).
T232 x CM37
Co159 X TX303
B. Burr’s BNL mapping work (1000+ markers)
Grown on St. Paul campus (2 years)
•Immortalized F2 (maintained by bulked F3 seed pool
to represent the F2)
Tx303 X Co159 IF2 (Ed Coe / Georgia Yerk-Davis)
Grown in St. Paul and at the University of Missouri
at Columbia (samples bulked).
13. Materials And Methods Cont.
•Endosperm collection:
10-24 DAP (2 day intervals)
Fix in farmers solution- ethanol:propionic acid (3:1)
Store in 70% ethanol
•Phenotypic traits: flowering time, kernel weight, endosperm cell
number, mean ploidy.
•Nuclei isolation: pectinase digestion for nuclei release
(phosphate buffer), RNase treatment, addition of propidium
iodide.
15. Materials And Methods Cont.
•Flow cytometry
•Coulter Epics MXL
•Agron ion laser (488 nm)
•Forward angle light scatter (FALS)- cell size qualities
•Right angle light scatter (RALS)- cell surface qualities
•Photomultiplier (fluorescence)
•Area (integrated or total fluorescence)
•Peak (AUX designation)
•Due to the large variation in DNA content, signals
were expressed with a logarithmic
transformation (Log) (FL3 Log-PI ).
•Data Analysis: ModFit LT 3.0
•Cell cycle modeling gaussian components are used to
model the “C” (DNA Content) Peaks- nonlinear
regression
16. •Qualitative Trait- Single recombinant on either side
of the gene defines location.
•Quantitative Trait- Recombination helps to define a support interval
but with the rest of the genome segregating and with influences of the
environment and epistastic interactions, the identification of precise
gene locations is difficult. LOD curve is used to show the most likely
genomic region.
Complex traits do not show perfect cosegregation with any single locus:
Polygenic Inheritance
(Lander and Schork, 1994)
Qualitative vs. Quantitative Genetics
17. Quantitative Trait Locus (QTL) Analysis Using
Molecular Markers in an RIL Population
P1 P2 RILs
Locus A: Associated with kernel size
Locus B: Not associated with kernel size
B1
B2
A1 A1 A2 A2
A1
A2
B1 B1 B2B2
Marker Class Means
37. Multi-year trait heritability estimates
T232 X CM37 RIL population
(Narrow sense- No dominance
variation in RIL)
Note: Dilkes et al. (2002) obtained a similar heritability estimate
(0.410.13) for the trait of mean ploidy (19 DAP) using a
parent-offspring regression approach (narrow sense).
38. Genetic correlations for select T232 X CM37 RIL
traits (St.Paul, MN 1996 and 1997). Multivariate
analysis (REML).
39. Genetic correlations for select T232 X CM37 RIL
traits (St.Paul, MN 1996 and 1997). Multivariate
analysis (REML).
41. Genetic map of all the endosperm cell number and
mean ploidy QTLs identified in the three mapping populations
42. Chromosome 7 in Detail
1. % Phenotypic
Variance explained
2. Additive Value
(in thousands)
3. LOD Score
Box Height = 3 LOD
Support Interval
43. IF2 endosperm
cell number
(Missouri)
Genome Scan
The multiple QTL model,
including all significant
regions simultaneously,
accounted for 35.0 ± 7.3%
of the total phenotypic
Variance.
44. Immortalized Tx303 X CO159 F2 Chromosome 7 for the trait
mean endosperm cell number at 16 DAP (Columbia, MO 1996).
Genetic Effect (a) 80×103± 28×103 cells (CO159 direction)
46. Summary of identified QTLs for the trait mean
endosperm nuclear ploidy.
AUC (area under curve) is derived by
integration between 14 to 24 DAP.
47. Candidate loci for endosperm cell
number and endoreduplication
(based on positional and biological
information)
Endosperm Cell Number
Endoreduplication
Sugar transport1 (stp1) chromosome 8 (bin 8.02-3)
reduced endosperm2 chromosome 7 (bin 7.03-4)
Basal endosperm transfer layer 1c: chromosome 2 (bin 2.08)
Zea agamous3 (ser/thr phosphatase): chromosome 4 (bin 4.06)
Zea agamous5 (ser/thr phosphatase): chromosome 5 (bin 5.06)
Soluble invertase2: chromosome 5 (bin 5.04)
48. Mitotic Repression
•Mitotic repression: protein and RNA synthesis disruption
during the mitotic cell cycle.
•Value of endocycle
(recognizing role of cell number, starch, etc.)
•Escape from mitotic repression coupled with
transcript abundance
•Rapid differentiation and intensive growth
•Even if the potential efficiency gained per cell is small,
when multiplied out by hundreds of thousands of cells,
as in the case of the maize endosperm,
a large effect on productivity could result.
49. Zea diploperennis Endosperm
Development
•Zea diploperennis is one of the four species in genus Zea.
•Perennial diploid teosinte (native to Mexico).
•Successfully crosses with Z. mays.
•Has pistillate spikes that bear 6-12 small kernels in hard casing.
•Short day treatment in Minnesota to induce flowering
•Does endoreduplication occur during Z. diploperennis
endosperm development?
•Use genetic variability of Z. diploperennis to expand
the genetic variability for cultivated maize kernel traits?
•John Doebley’s cultivated maize X Z. diploperennis
mapping population would be a valuable resource to
investigate this issue.
50. Endosperm Development
Zea diploperennis
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Liquid Endosperm Gelatinous Solid
Pedrosa and Vasil,
Maydica 41 (1996) 333-348
Days after pollination
54. Challenges
•Study cytological traits at the whole plant level
•Account for kernel number per ear, ears per plant,
plant density
•Develop near-isogenic lines to identify novel genes
•Expense of phenotyping cytological traits at the
population level
55. Future Directions
•Investigate hexose/sucrose ratio during endosperm development
Metabolic control of development / invertase control theory
Hexose sugar as signaling molecule for cell cycle
High hexose ratio (mitosis promoting factor)
Low hexose ratio (differentiation signal, endocycles)
Cotyledon tissue (Vicia faba)
(Wobus and Weber, Biol. Chem., Vol. 380, pp. 937-944.)
•Investigate hormonal control of endocycle
(Phytohormone X sugar status interaction, for example)
•Transgenic modification of key cell cycle regulation genes
ex. Cyclin-dependent kinase A
(Dr. Brian Larkin’s Group at Arizona)
•Genomic/microarray analysis of the developing tissue along a
time-course to identify particular genes that benefit from endocycle.
56. Conclusions
•Natural genetic variation identified for both
endosperm cell number and extent of endoreduplication
•Efficient estimation of 3-96C class peaks by the flow
cytometry data analysis software MODFIT LT 3.0
compared to manual vertical integration methods
•Broad sense heritability estimates
Endosperm cell number trait (ranged 0.16 to 0.56)
Mean ploidy trait (ranged from 0.14 to 0.77)
•Genetic correlation between mean ploidy and endosperm
cell number:
Weighted average (all populations, two years): -0.27
•Genetic correlation between MTC and kernel weight
0.65 (0.46) in 1996 and 0.67 (0.66) in 1997
Conclusions continued on the next slide
57. Conclusions cont.
•QTL identification
•Endosperm cell number
•8 genomic regions identified
•Range of effect (2a) 138 × 103 to 312 × 103
cells.
•Extent of endoreduplication
•10 genomic regions identified
•Range of effect (2a) 0.96 C to 2.38 C mean
ploidy units
•Endoreduplication was detected in endosperm samples
collected from both Zea diploperennis and Tripsacum
dactyloides
58. Acknowledgements
Ronald Phillips
Friedrich Srienc
NIH Biotechnology Traineeship Program
Richard Kowles
Georgia Yerk-Davis
Robert Jones
Jeff Roessler
Larry Carlson
Suzanne Livingston
Jayanti Suresh
Cristian Vladutu
Mike Olsen
