Pioneer Hi-Bred International
University of Missouri
bk3, a new brittle stalk mutant of maize --Multani, DS, Johal, G A brittle stalk mutant was identified from the self-pollinated population of a plant known to have Mutator activity in 1997 (MO97-08-626). The true breeding nature of the mutant was confirmed by growing self-pollinated progeny of the WT-sib. Leaves of this mutant are highly brittle and snapped readily when pressed between fingers (Fig. 1). All other parts of the plant, such as the stalk and roots, are also brittle and easily breakable. In addition, this mutant is weak, dwarf and has leaves that develop a reddish tinge especially towards at the tip. The inheritance of this mutant was studied by crossing with the inbred B73. The F1 was normal in phenotype and in an F2 population of 108 plants, 82 were normal and 26 were mutant. These data agree with a 3 normal: 1 mutant ratio. Thus, the new brittle stalk mutant phenotype is due to a single recessive gene. To determine its allelic relationship with another known brittle stalk mutant, bk2 (Langham, MNL 14:21-22, 1940), the two were crossed (bk2 was acquired from the Maize Genetics Coop). The resulting F1 was normal in phenotype; the appearance of both brittle stalk mutants in F2 indicates that the new brittle stalk mutant is non-allelic to bk2.

Many features of this new mutant resemble those described for bk1 (Brewbaker, MNL 69:58-59, 1995). However, because of the unavailability of seed for bk1, which appears to have been lost from the Maize Genetics Coop, allelic relationships between the two can never be established. Therefore, the new brittle stalk mutant has been designated as bk3.

It is difficult to maintain this mutant as a homozygote, and only occasionally has it been seen to shed pollen. Thus, it is maintained as a heterozygote. In an attempt to map bk3 to the specific chromosome arm, we used B-A translocations. Plants heterozygous for bk3 (bk3/+) were used as females and crossed with TB-stocks (received from Jim Birchler, University of Missouri-Columbia) representing 16 (TB-1Sb, TB-1La, TB-2Sa, TB-3Sb, TB-3La, TB-4Sa, TB-4Lb, TB-5Sc, TB-5Lb, TB-6Lc, TB-7Sc, TB-7Lb, TB-8Lc, TB-9Sd, TB-9Lc, TB-10Sc, and TB-10L19) out of 20 chromosome arms. Except for TB-9Lc, progeny of all crosses was normal in phenotype. In the case of the cross with TB-9Lc, 11 out of 126 progeny plants tested were mutant, suggesting that bk3 is located on the long arm of chromosome 9. Interestingly, bk2 also maps to 9L (Langham, 1940).

As bk3 was isolated from a Mutator population, a segregating F2 population was analyzed by co-segregation analysis to identify the Mu1 element that segregated completely with the mutant phenotype. This Mu1 element, along with the flanking genomic DNA (5.4 kb/NotI fragment), was cloned and subject to a higher resolution of linkage with the mutant allele. One out of 96 plants tested gave us an exception to the tight linkage between the clone and the mutant phenotype, indicating that the clone we have isolated does not come from the bk3 locus. However, the two are localized within 1-2 cM of each other. This information was used to reconfirm the chromosomal location of bk3 using the collection of oat-maize addition lines. Two gene specific primers from this bk3-linked clone were designed and used to amplify the target sequence from DNA derived from each of the ten oat-maize addition lines. An amplification product, which hybridized positively with a clone-specific probe, was obtained only from the oat-maize addition line 9 (Fig. 2). This result, along with data from the B-A study, clearly established that like bk2, bk3 is localized to 9L.

Figure 1. bk3, brittle stalk mutant: brittle plant parts after 4-leaf stage are easily broken. Lower leaves also have reddish tinge more at the tip.

Figure 2. Chromosomal location of DNA fragment showing tight linkage with bk3 in corn: a) PCR amplification using oat-maize addition lines DNA; b) Southern blot of PCR products.

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