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A study was made of the genetics of resistance to the soybean …


Biology Articles » Genetics » Ecological Genetics » Genetics of Resistance to Soybean Cyst Nematode, Heterodera glycines Ichinohe (RACE 3), in a Brazilian Soybean Population » Materials and Methods

Materials and Methods
- Genetics of Resistance to Soybean Cyst Nematode, Heterodera glycines Ichinohe (RACE 3), in a Brazilian Soybean Population

To develop the population for this study, crosses were made between the soybean genotypes BR 90-4722 and FT-Cristalina. These genotypes are, respectively, resistant and susceptible to SCN, race 3 (Arantes, 1997). Part of the resulting F1 seeds were kept in a cold room and part was used to obtain the F2 seeds. Parents (resistant and susceptible) along with their F1 and F2 generations were sowed in boxes containing sterilized sand as a substrate. When the seedlings had completely expanded cotyledonar leaves, they were transferred to three-liter pots containing a 2:1 soil:sand mixture previously sterilized.

The inoculum of H. glycines (race 3) was obtained from fields infested with SCN, race 3, in the county of Iraí de Minas. The race of the nematode was confirmed through tests in differential soybean cultivars performed according to suggestions of Arantes (1997). Thirty-five-day-old plants were collected from these fields and the females were extracted from the roots and macerated in a system of sieves. Two days after transferring the seedlings to the pots, the inoculation was made using a suspension containing approximately 11,000 eggs and juveniles. The number of eggs and second stage juveniles was obtained by counting them on a Peters plate under an optical microscope.

The suspension was applied near the stem of each seedling. Plants were irrigated after the inoculation to avoid the drying out of the nematodes. To evaluate the degree of resistance of each individual, the plants were carefully taken from the pots 34 days later, and the number of females and cysts present in the roots of the parental, F1 and F2 generations was obtained by counting them.

To study the genetics of the resistance of the population to SCN, the number of cysts found on each plant of each generation was first considered. The means, standard errors, variances and expected values for the generations F1 and F2 were estimated according to the procedures employed by Mauro et al. (1995). Thus, the means, the variances and the standard errors for each generation were estimated as follows: means = µ = ∑fixi/∑fi; variances = σ2 = (∑fixi2 - ∑fiµ2)/∑fi; standard error = (σ/n)1/2; where: fi = frequencies; xi = observed values and n = number of individuals.

The expected means (E) for the generations F1 and F2, considering an additive model, were estimated as follows: E(F1) = (µP1 + µP2)/2; E(F2) = [F1 + E(F1)]/2. These estimates were compared with the observed values for each generation through the t-test for independent samples with unequal variances, as proposed by Snedecor and Cochran (1989). Comparisons between the observed values for the F1 and F2 generations and between the F1 and the resistant parent were also performed. To verify the significance of these comparisons, the calculated t was obtained as follow063004m01.gif where µ1 and µ2 = observed and expected means;063004m02.gif and063004m03.gif = estimated variances, and n1 and n2 = size of the samples. The number of degrees of freedom associated with each one of the comparisons was obtained through the expression:063004m04.gif where v1 = n1 - 1 and v2 = n2 -1; V1 = 063004m05.gif/n1 and V2 =063004m06.gif/n2.

The number of genes related to the resistance of the population to the SCN, race 3, was determined by using the following expression:063004m07.gif  where n = number of genes related to the character; µP1 = mean for parent 1; µP2= mean for parent 2; 063004m08.gif = variance for the generation F1 and sF2 = variance for the generation F2. The heritability for the resistance to SCN in the same population was also estimated, being used the methodology proposed by Mahmud and Kramer (1951). This methodology estimates the heritability in the broad sense, which is given by: 063004m09.gif where:063004m10.gif  = variance for parent 1;063004m11.gif= variance for parent; 2063004m12.gif = variance for generation F1, and063004m13.gif = variance for generation F2.

To study the type of gene action and to confirm the number of genes related to the resistance of the population to SCN, a chi-square test, as described by Snedecor and Cochran (1989), was performed. The calculated value for the chi-square test was given by:  α2 = ∑[(o - e)2 / e], where o and e are, respectively, the observed and the expected frequencies. For this analysis, plants from the parents, F1 and segregant population were classified according to their reaction to the inoculation with SCN, using the rating system proposed by Hartwig (1985), as follows: 0 - absence of females and cysts; 1 - from 1 to 5 females and cysts; 2 - from 6 to 10 females and cysts; 3 - from 11 to 20 females and cysts; 4 - more than 20 females and cysts. Plants scored from 2 to 4 were considered as susceptible and the cultivar FT-Cristalina was used as a standard of susceptibility, since according to Arantes (1997) it can substitute Lee 68, with the advantage of being a Brazilian soybean cultivar.


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