Download Ch. 5: Presentation Slides

Contents
DAY 1: Molecular biology, the basics
• Chapter 1: DNA structure and gene
expression, nuclear and organel genomes 53
• Chapter 2: DNA replication, PCR,
electrophoresis, DNA sequence analysis 33
• Chapter 3: Organel and ITS sequence, DNA
databases, phylogenetic analysis 46
• Pract.: DNA BLAST, alignment, phylogeny
Molecular techniques in plant breeding, Cantho september 2006
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Contents
DAY 2: Methods for the analysis of DNA
polymorphisms
• Chapter 4: Recombinant DNA, restriction
enzymes, cloning, transformation 21
• Chapter 5: Hybridisation, RFLP, micro-array
& SNP analysis 33
• Chapter 6: PCR methods for analysis of
DNA polymorphisms 29
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Contents
DAY 4: Marker assisted breeding
• Feedback on molecular techniques
• Chapter 7: The use of molecular markers in
breeding 18
• Chapter 8: Fingerprints, diversity analysis,
specific markers 19
• Chapter 9: Genetic linkage, genetic maps,
Introgression, QTL analysis 42
• Pract.: Demonstration AFLP, SSR, QTL
analysis,
genetic
Molecular
techniques inmaps
plant breeding, Cantho september 2006
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Contents
DAY 5: Functional genomics, transformation
technology
• Chapter 10: Plant transformation technology 29
• Chapter 11: Functional genomics 11
• Chapter 12: Transgenic crop plants 31
• Examination
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Chapter 1: Genes and genomes
DNA structure
Genomes
Protein structure
Gene structure and expression
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Cells and DNA
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Chromosomes and DNA
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DNA: Chemical Composition
• Two types of nitrogen-containing bases
comprise the chemical structure of DNA:
- purines = adenine and guanine, A & G
- pyrimidines = thymine and cytosine, T & C
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DNA: Chemical Composition
• Nucleotides = building blocks of DNA =
phosphate + sugar + base
• Nucleoside = sugar +base
• Sugar = 5 carbon
deoxyribose
• Phosphodiester bonds
link sugar molecules
to phosphate groups
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DNA: Chemical Structure
• DNA = polynucleotide = a chain of bases
• Orientation of sugar-phosphate linkages =
5’ to 3’ as the phosphate
attached to the 5’ carbon
of one sugar is linked to
the 3’ carbon of the next
sugar
• Purine and pyrimidine
bases are linked to the
1’ carbon of sugar
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This polarity is extremely important!!!!
5’ end:
P
3’ end:
OH
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DNA: Chemical Structure
• DNA consists of two polynucleotide chains
which run 5’ to 3’ in opposite
directions = antiparallel
• DNA chains are held together by
hydrogen bonds between bases
• DNA bases pair by Chargaff’s
rules:
- Adenine (A) pairs with
Thymine (T) by 2 H-bounds
- Guanine (G) pairs with
Cytosine
(C) by 3 bounds
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3’ end:
5’ end:
5’ end:
3’ end:
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DNA Sequence: convention 5’ to 3’end, one strand
(because other strand is complementary and therefore known also)
5’
3’
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Genome Size
• Complex organisms have large
genomes = genetic contents of a cell
• Genomic size increases with evolutionary
complexity (in general!)
• Size of DNA is measured in kb = kilobase
pairs
• Size of large genomes is measured in
Mb = megabase pairs
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Genes and genomes
• Gene = unique sequence of DNA bases, coding
for a protein
• Alleles = different variants of a gene in different
organisms or on the homologous chromosomes
Organism
Genome
size
Gene
number
E. coli
4.6 Mb
2800
Yeast
12 Mb
6000
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Genes and genomes
Organism
Genome
size
Gene
number
C. elegans
100 Mb
20,000
A. thaliana
120 Mb
26,000
H. sapiens
3,000 Mb
40,000?
Z. mays
2,500 Mb
26,000?
Salamander
90,000 Mb 30,000?
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Genes and genomes
Eukaryotic genomes contain much DNA outside the genes
Human
Yeast
Z. mays
E. coli
Genomes 2
T.A. Brown, Bios
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Chromosomes of a human being:
2 copies of each chromosome,
ordered by size on a slide
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Chromosomes and DNA
Eukaryotes are diploid
(2 homologous chromosomes,
with alleles of genes)
Eukaryotes have linear
chromosomes in their nucleus
This picture shows a duplicated
chromosome= 2C = 2 identical
chromatids, the scheme
unravels one of those
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Genomes
nuclear DNA:
From both parents
(biparental)
Chloroplast (cp) and
mitochondrial (mt)
DNA:
Usually maternal
Plant cell
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Genomes
• Prokaryotes have a circular genome and
often also small circles = plasmids
• Mitochondria (and chloroplasts) in
eukaryotic cells have their own genome
= circular (endosymbiont hypothesis)
Prokaryote was swallowed by an early eukaryote and
degenerated, it now provides energy (mitochondria)
or performs photosynthesis (chloroplast)
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Mitochondrial genomes
•
•
•
•
Human: 16 kb
Yeast: 84 kb
Corn: 570 kb
Genes for:
rRNA, tRNA
proteins for
ATP synthesis
yeast
human
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Chloroplast genomes
• 120 – 200 kb
• Genes voor:
rRNA and tRNA
RNA polymerase
ca. 120 proteins
for
photosynthesis
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Maternal inheritance of mitochondrial DNA
Maternal inheritance through egg cytoplasm, no mixing,
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in plant breeding,
Cantho september
2006
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no recombination:
excellent
for lineage
analysis
Organel DNA for tracing back
Mystery of Russian tsar-family: bones in grave could be
identified based on comparison of mitochondrial DNA
with that of living descendants (through the maternal
line). Many copies (many circles per organel, many
organels perMolecular
cell)techniques
of thein plant
DNA
allow
breeding,
Canthoefficient
september 2006 analysis.27