Waxy wheat starch types having waxy proteins in granule

Details Waxy wheat starch types having waxy proteins in granule Waxy wheat starch types having waxy proteins in granule

1999-04-08

2002-03-26

Leary, Louise N. (Department: 1623)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

C536S045000, C536S102000, C800S320300, C800S320000, C800S298000, C800S260000, C800S263000, C800S276000, C426S018000, C426S549000

Reexamination Certificate

active

06361935

ABSTRACT:

FIELD OF THE INVENTION
Broadly, the present invention relates to mutating starch genes in polyploid cereal grains. Specifically, this invention concerns mutant wheat plants, mutant wheat grain and the starch therefrom.
BACKGROUND
Cereal grains, such as rice, wheat, corn and barley help to feed the world's population. A large percentage of the world population's staples are formed of these grains. These grains are processed to make breads, cereals, pasta, flour, etc. Processed grains have different qualities which lead to different product uses. Wheat grains are processed into wheat flour, which is a storehouse of nutrients. The starch, protein, lipids, enzymes, and nutrients affect, in differing degrees, the flour product. Starch in the flour product affects its characteristics to a large degree. The digestibility, processing temperatures, cooking qualities of flour are all impacted by the type of starch used.
Starch is formed of two components amylose and amylopectin. At least one of these components is altered in a number of diploid cereal grains which have starch mutations. One starch mutation is referred to as the waxy mutation. Cereal grains having the waxy mutation form low amylose starch. Naturally occurring waxy mutants are well known in rice and maize, both diploid species. However, naturally occurring starch mutants are not known in polyploid species. To alter the starch in a polyploid species requires several independent mutations. Wheat seldom has naturally occurring mutants because both soft and hard wheats are hexaploid, although durum wheats are tetraploid. No one has discovered a naturally occurring waxy wheat.
Wheat (
Triticum aestivum
L.) has three chromosome sets derived from three different species. Each chromosome set has a genome letter, A, B or D. A wheat waxy mutant would have homozygous waxy alleles in each of the A, B & D chromosomes. Mutations in wheat have been identified by protein characterization in the individual A, B & D genomes. Until 1992 these protein characterizations detected only a single waxy protein band. The test was not able to distinguish a protein band for each of the three genomes. If one of the three genomes was not producing a protein this test could not detect it. A modified detection system using SDS-PAGE with low BIS acrylamide concentration and a two-dimensional gel electrophoresis (2-D PAGE) showed two of the expected three protein bands. The third band was detected by using isoelectric focusing (IEF) for the first dimension and the modified SDS-PAGE for the second. The modified detection system detected three protein bands. Each protein band corresponds to one of the A, B and D sets of chromosomes. By detecting the individual proteins wheat lines could be screened for null waxy alleles. A null allele does not produce a certain protein at that allele on a certain chromosome. A null mutant does not produce a certain protein at any of the chromosomes. This is in contrast to a non-null mutant which does produce the protein, but in an inactive state.
Once a test showing the three bands was identified a number of researchers began to screen waxy wheat for null alleles. Single null alleles were located when individual starch proteins were missing from wheat starch. Single null waxy wheat alleles were identified in only about 10% of the US winter wheat germplasm. The remaining wheat were wildtype having three functional wx loci. R. A. Graybosch reported on a few single null waxy alleles in both the A and B genomes. Only two single nulls are known to exist in the D genome. One single null D genome waxy allele has been reported in Japan and one has been reported in Canada.
Recently, researchers discovered four separate double null waxy alleles in wheat. Each of these double null waxy alleles (partial mutants) were null for waxy alleles of the A and B genome. The Japanese reported the Kanto lines 79, 107, Saikai 173 and R. A. Graybosch reported Ike. Ike is a public line, developed by the Kansas University breeding program. These double null waxy partial mutants are the only ones known to exist. However, with the modified screening procedure researchers can expect the discovery of additional single and double null alleles. Like single waxy nulls, these newly discovered double null waxy partial mutants, still produce a waxy protein in the D allele and thus still have significant amylose content. However, the double null waxy partial mutants amylose content is recognizably less then single waxy nulls.
Even after the discovery of the double nulls there still was needed a waxy wheat mutant plant that had a waxy mutation in all three genomes. In 1994, a waxy wheat plant free of waxy protein was produced. The Bai Huo Chinese cultivar lacking a waxy protein was crossed on to Kanto 107 and Saikia 173. Out of 720 F2 seeds 14 were free of Wx proteins.
Conventional crossing of a partial null mutant with a single null allele did produce a waxy wheat. It also produced an entirely new combination of genes within the chromosomes of the resultant waxy wheat. To generate useful lines from the resultant waxy wheat plant breeding was used. The waxy wheat was selfed and the progeny were selected for agronomics traits and the waxy trait throughout breeding generations. Although conventionally bred plants having the waxy mutation are easily identified by the iodine test; agronomic traits are much more difficult to identify. Agronomic traits are often multigenetic and in wheat these are further complicated by three separate sets of chromosomes. Reconstruction of the three chromosomes with conventional breeding, or even with dihaploidy takes time and a number of crosses. Even after a number of crosses, the waxy wheat is not essentially identical to one parent, it is only similar to the parent.
A plant which is essentially identical to the parent plant is an isogenic line. An isogenic line is characterized by essentially identical genes. Forming a waxy wheat that is isogenic to its parent avoids conventional breeding problems. There is a need for a method of forming a waxy wheat that does not produce an entirely new combination of genes within the chromosomes of the waxy wheat. There is a need for an efficient method for forming full mutants from double chromosome mutants or double chromosome mutants, from single chromosome mutants in polyploid cereals. In other words, there is a need for a method to form isogenic polyploid lines which contain mutations.
Additional needs develop, once an isogenic starch mutation is in commerce. Breeders will breed with this isogenic germplasm to place the starch mutation into other germplasm. To maintain germplasm security, there is a need for a method of identifying the mutation. If the mutation is identified then misuse of germplasms can be identified.
SUMMARY OF THE INVENTION
An object of the present invention is a method of forming polyploid isogenic seeds and plants which differs from the parent by being a starch mutant.
Still another object of the present invention is an isogenic wheat seed and plant that produces waxy starch.
A further object of the present invention is a method of identifying isogenic lines by fingerprinting the modified starch the lines produce.
Yet a further object of the present invention is waxy wheat starch which contains an inactive protein in at least one of the waxy loci.
In addition, another object of the present invention is the method of mutating polyploid cereals to form a modified starch.
The present invention broadly encompasses a method of making a full mutant allele polyploidy cereal seed. This method includes the following steps. Treating a double mutant allele in a polyploid plant material with a mutagen. This forms treated plant material. The treated plant material is then screened to identify full mutant allele plant material. The full mutant allele plant material are selected. The present method can also include the additional steps of selecting polyploid plant material having a single mutant allele and treating this plant material with a mutagen then screening the plan

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