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Mutation - Mutant
Mutation
Changes in genes and chromosomes
Mutated
Altered genes
Mutant
New organism with a mutated gene or rearranged chromosomes
Mutation Breeding
Advantages
Screen very high populations (cell based) Can apply selection to single cells
Disadvantages
Many mutations are non-heritable non Requires dominant mutation (or double recessive mutation); most mutations are recessive
Can avoid this constraint by not applying selection pressure in culture, but you loose the advantage of high through-put throughscreening have to grow out all regenerated plants, produce seed, and evaluate the M2 Alternative: perform on haploid cell lines
Mutation
May involve any trait All kind of transition are encountered, from drastic morphological changes deviations in physiology so minute as to be almost indiscernible Harmful or even lethal
Type of mutation
Spontaneous (natural) mutation
Some have played an outstanding role in development of valuable crop cultivars and hybrids 2. Unfortunately, it can not form the basis of modern plant breeding due to its low frequency and difficulties in detection 1.
Induced mutation
Chromosomal mutation
Mutation associated with splitting and subsequent changes in the structure of the chromosomes The end of the split chromosomes may fuse to form structure again, but the new chromosomes are not always exactly what the used to be The microscopic structures of chromosomes may be characterized by deletion or deficiency (loss of a chromosomal segment), duplication (doubling of a chromosomal segment), inversion (rearrangement of a group of genes in a chromosomal segment in a such a way that their order is reversed; rearrangement of genetic material in a chromosome results from loss of segment, its rotation by 180, and fusion of the separated ends) and 180 translocation (change in a position of a chromosome or more often exchange of segments between different chromosomes)
Genome mutation
Changes in sets of chromosomes
Remarks: 1. Breeders are more interested in gene mutation, because chromosomal rearrangement usually produce negative results, such as lower fertility of the offspring 2. Mutant are aften of great value for breeding as sources of new, previously unknown useful characters 3. Mutagenesis may be instrumental in obviating the technical difficulties arising in the crossing of such a small flowered crops such as milled
Physical mutagens
4. The object can be irradiated in two ways:
5. 6.
With an aid of a powerful source of a short-duration gamma shortrays for short duration radiation. Need special units for irradiating living object A much weaker radiation but operating continuously (gamma field). the dosage must be varied depending not only on the plant species whose seeds/organs are irradiated, but also on many other factors plant must be irradiated heavily enough to ensure as many inherited changes as possible but without seriously affecting the germination, growth and fertility of plant directly emerging from the irradiated seeds or vegetative organs (critical (critical radiation dose: dosage which strong enough to assure many dose: mutation not yet so strong as to kill plants) plants)
Chemical mutagens
Mutagenic substances belonging to different classes of chemical compounds, such as ethylene imine, diethyl sulfate, dimethyl sulfate, N-nitrosoethyl urea, NNNnitrosomethyl urea, methal sulfonate, diepoxy butane, ethyleneoxide Most are highly toxic, usually result in point mutations Use in solution in the concentration ranging from tenth hundredths even thousandths of percent Many chemical mutagens are much more effective than physical one. If irradiation of crops produces 10 15% of viable inherited changes, chemical mutants do the same at a rate of 30 to 60% They often exert more specific and finely tuned action on the cell
Chemical mutagens
Some substances (supermutagen) are capable of causing inherited changes in plants at a rate up 100% Chemical mutagens aim at the most vulnerable spot of a living organism (DNA) to induce changes in nucleotides and alter the genetic information (Sometimes causes specific mutation) It provides a powerful tool to induce desire changes in a trait
It has been much easier and quicker to obtain variant plant types Specific location of the mutation event (segmental chimera) becomes important. The mutant must be in meristematic tissue that will produce faithfully through cutting or other vegetative means Bud Scion Cutting Tuber bulbs
Clear Objective
Can t expect to just plant things out and see what happens; relates to having an effective screen This may be why so many early experiments failed
Mutation detection
Detection, isolation and testing mutants are extremely difficult Due to the sporadic nature of viable useful mutations, it is advisable to have larger plant population When mutagens are used in breeding, the biological nature of the trait (dominance or recession of the mutation) and crops must be taken into account
1. 2. 3. 4.
Theoretically possible for any enzyme-targeted herbicide it s enzymerelatively easy to change a single enzyme by changing a single gene
Mutagenesis - Overview
M3 plant mut-2/mut-2 Case 1:
x
Legend
M3 plant mut-1/mut-1 Case 2:
x
M3 plant mut-1/mut-1 Wild type +/+
D Backcrossing
A Mutagenesis
(chemical, radiation, T-DNA,) Mutant phenotype BC1 plant mut-1/+ Multiple backcrosses to remove background mutations
x
mut-1/mut-2 mut-1/+ mut-2/+ No allelism Allelism Single gene Two genes Wild type +/+
M1 plants Harvested In pools Pool 1 Pool 2 Pool 3, etc. Wild-type phenotype Strain A/A Pools of M2 seeds Strain A/B Strain B
C Allelism Tests
How many genes are involved?
Recessive phenotypes appear here F Mapping Screen M2 pools (1, 2, etc.) +/+ for mutant phenotypes Where is the gene located? Wild type +/+ Strain B Propagate mutant from mut1/mut-1 or from its mut-1/+ heterozygous siblings mut-1/mut-1 Option 1: Backcross
Outcross
x
mut-1/mut-1 Strain A
mut-1/mut-1
B Screening
G Gene Cloning
mut-1/+ Strain A/Strain B How does the gene function?
x
mut-1 Strain A
M3 seedlings
Re-screening Establish segregation ratio - Recessive or dominant? - Monogenic or polygenic? - Penetrance? Initiate mutant characterization
Option 2: Selfing
Examine co-segregation of mutant phenotype versus strain-specific (visible or molecular) traits Mapping population Mapping population
A Mutagenesis
Legend
Pool 1
Pool 2
Pool 3, etc.
Pools of M2 seeds
Wild-type phenotype
B Screening
Screen M2 pools (1, 2, etc.) for mutant phenotypes M2 seedlings Mutant mut-1 Propagate mutant from mut1/mut-1 or from its mut-1/+ heterozygous siblings
Legend
Mutant phenotype
M3 seedlings
Re-screening Establish segregation ratio - Recessive or dominant? - Monogenic or polygenic? - Penetrance? Initiate mutant characterization
Wild-type phenotype
C Allelism Tests
x
M3 plant mut-2/mut-2 Case 1: M3 plant mut-1/mut-1 Case 2:
Legend
Mutant phenotype
mut-1/mut-2
Wild-type phenotype
D Backcrossing
M3 plant mut-1/mut-1 x/x
F Mapping
Where is the gene located? +/+ Outcross mut-1/mut-1
mut-1/mut-1 Strain A
G Gene Cloning
How does the gene function?
x
Strain A/A Strain A/B Strain B mut-1 Strain A
Option 2: Selfing