Cláudio Lopes de Souza

Comparison of RAPD, RFLP, AFLP and SSR markers for diversity studies in tropical maize inbred lines

In order to compare their relative efficiencies as markers and to find the most suitable marker for maize diversity studies we evaluated 18 inbred tropical maize lines using a number of different loci as markers. The loci used were: 774 amplified fragment length polymorphisms (AFLPs); 262 random amplified polymorphic DNAs (RAPDs); 185 restriction fragment length polymorphisms (RFLPs); and 68 simple sequence repeats (SSR). For estimating genetic distance the AFLP and RFLP markers gave the most correlated results, with a correlation coefficient of r = 0.87. Bootstrap analysis were used to evaluate the number of loci for the markers and the coefficients of variation (CV) revealed a skewed distribution. The dominant markers (AFLP and RAPD) had small CV values indicating a skewed distribution while the codominant markers gave high CV values. The use of maximum values of genetic distance CVs within each sample size was efficient in determining the number of loci needed to obtain a maximum CV of 10%. The number of RFLP and AFLP loci used was enough to give CV values of below 5%, while the SSRs and RAPD loci gave higher CV values. Except for the RAPD markers, all the markers correlated genetic distance with single cross performance and heterosis which showed that they could be useful in predicting single cross performance and heterosis in intrapopulation crosses for broad-based populations. Our results indicate that AFLP seemed to be the best-suited molecular assay for fingerprinting and assessing genetic relationships among tropical maize inbred lines with high accuracy.

Comparison of similarity coefficients used for cluster analysis with dominant markers in maize (Zea mays L)

The objective of this study was to evaluate whether different similarity coefficients used with dominant markers can influence the results of cluster analysis, using eighteen inbred lines of maize from two different populations, BR-105 and BR-106. These were analyzed by AFLP and RAPD markers and eight similarity coefficients were calculated: Jaccard, Sorensen-Dice, Anderberg, Ochiai, Simple-matching, Rogers and Tanimoto, Ochiai II and Russel and Rao. The similarity matrices obtained were compared by the Spearman correlation, cluster analysis with dendrograms (UPGMA, WPGMA, Single Linkage, Complete Linkage and Neighbour-Joining methods), the consensus fork index between all pairs of dendrograms, groups obtained through the Tocher optimization procedure and projection efficiency in a two-dimensional space. The results showed that for almost all methodologies and marker systems, the Jaccard, Sorensen-Dice, Anderberg and Ochiai coefficient showed close results, due to the fact that all of them exclude negative co-occurrences. Significant alterations in the results for the Simple Matching, Rogers and Tanimoto, and Ochiai II coefficients were not observed either, probably due to the fact that they all include negative co-occurrences. The Russel and Rao coefficient presented very different results from the others in almost all the cases studied and should not be used, because it excludes the negative co-occurrences in the numerator and includes them in the denominator of their expression. Due to the fact that the negative co-occurrences do not necessarily mean that the regions of the DNA are identical, the use of coefficients that do not include negative co-occurrences was suggested.

Combining ability of inbred lines of maize and stability of their respective single-crosses

The utilization of diallel crosses for identification of superior combinations is a common practice in maize (Zea mays L.) breeding programs. This methodology allows the estimation of the combining ability of genotypes being evaluated. In this work, five inbred lines were evaluated as to their general (GCA) and specific (SCA) combining abilities, by using a complete diallel scheme. The single-crosses produced between these inbred lines were evaluated in seven environments, along with two checks, by using a randomized complete block design. Traits analized were: grain yield, plant height, ear height, ear placement, and prolificacy. A diallel analysis was carried out, following an adaptation of Griffings method IV, in addition to hybrid stability and adaptability analyses. Significant differences were detected for entries and environments for all traits. The interaction genotype vs. environment was significant for all traits. GCAs were significant for all traits, while SCAs were non-significant only for ear placement. For grain yield, both additive (GCA) and non-additive (SCA) effects were important, while for the remaining traits additive effects were more important. The high yielding single-cross was obtained from the cross of lines L-08-05F and L-38-05D. Those inbred lines showed higher GCAs and their cross also had high SCA; also, it is responsive to environment improvements and reasonably stable. The second in rank high yielding single-cross, L-46-10D x L-08-05F, showed wide adaptability and stability.

Microsatellite-assisted backcross selection in maize

Abstract A microsatellite marker (SSR) was chosen to simulate a target allele and three criteria (02, 04 and 06 markers perchromosome) were tested to evaluate the most efficient parameters for performing marker-assisted backcross(MAB) selection. We used 53 polymorphic SSRs to genotype 186 BC 1 maize (Zea mays L.) plants produced bycrossing the inbred maize lines L-08-05 (donor parent) and L-14-4B (recurrent parent). The second backcross (BC 2 )generation was produced with 180 plants and screened with markers which were not recovered from the firstbackcross (BC 1 ) generation. A total of 480 plants were evaluated in the third backcross (BC 3 ) generation from which48 plants were selected for parental genotype recovery. Recurrent genotype recovery averages in three backcrossgenerations were compatible with those expected in BC 4 or BC 5 , indicating genetic gain due to the marker-assistedbackcrossing. The target marker (polymorphic microsatellite PHI037) was efficiently transferred. Six markers perchromosome showed a high level of precision for parental estimates at different levels of maize genome saturationand donor alleles were not present in the selected recovered pure lines. Phenotypically, the plants chosen based onthis criterion (06 markers per chromosome) were closer to the recurrent parent than any other selected by othercriteria (02 or 04 markers per chromosome). This approach allowed the understanding that six microsatellites perchromosome is a more efficient parameter than 02 and 04 markers per chromosome for deriving a marker-assistedbackcross (MAB) experiment in three backcross generations.

Reciprocal recurrent selection effects on the genetic structure of tropical maize populations assessed at microsatellite loci

A modified reciprocal recurrent selection (RRS) method, which employed one cycle of high-intensity selection, was applied to two tropical maize (Zea mays L.) populations, BR-105 and BR-106, originating the improved synthetics IG-3 and IG-4, respectively. In the present study the effects of this kind of selection on the genetic structure of these populations and their synthetics were investigated at 30 microsatellite (SSR) loci. A total of 125 alleles were revealed. A reduction in the number of alleles was observed after selection, as well as changes in allele frequencies. In nearly 13% (BR-105) and 7% (BR-106) of the loci evaluated, the changes in allele frequencies were not explained, exclusively due to the effects of genetic drift. The effective population sizes estimated for the synthetics using 30 SSR loci were similar to those theoretically expected after selection. The genetic differentiation (GST) between the synthetics increased to 77% compared with the original populations. The estimated RST values, a genetic differentiation measure proper for microsatellite data, were similar to those obtained for GST. Despite the high level of selection applied, the total gene diversity found in the synthetics allows them to be used in a new RRS cycle.

Responses to reciprocal recurrent selection and changes in genetic variability in IG-1 and IG-2 maize populations

This paper reports the effects of three cycles of reciprocal recurrent selection (RRS) on the means, genetic variances, and on the genetic correlations for several traits in the IG-1 and IG-2 maize (Zea mays L.) populations. Interpopulation full-sib progenies from cycle zero (C0) and from cycle 3 (C3) of RRS were evaluated in two locations. RRS was highly effective to improve the traits according the objectives of the program: grain yield and prolificacy increased significantly, while plant height, ear height, and ear placement decreased significantly. Genetic variances for all traits decreased significantly from C0 to C3, but the genetic correlations did not change consistently across the cycles of selection. The expected responses to the fourth cycle of RRS and the probability of selecting double-crosses from C3 that outperform those from C0 showed that the decreases in the genetic variances were not great enough to limit the continued improvement of the populations as well as the use of the improved populations as sources of inbred lines to develop commercial hybrids. However, if the magnitudes of the genetic variances continue to decrease, new sources of improved germplasm should be incorporated into both populations to allow the continued improvement of the interpopulation by RRS.

Response to recurrent selection under small effective population size

A formula was derived for the prediction of the response to recurrent selection when the effective population size (Ne) is small. Usually, responses to selection have been estimated by Rs = ics2A/sPh, where i, c, s2A, and sPh stand for standardized selection differential, parental control, additive variance, and phenotypic standard deviation, respectively. This expression, however, was derived under the assumption of infinite population size. By introducing the effects of finite population size, the expression derived was Rs = [ic(s2A + DFD1)/sPh] - DFID, where DF, ID and D1 are the changes in the inbreeding coefficient, the inbreeding depression, and the covariance of additive and homozygous dominance effects, respectively. Thus, the predicted responses to selection based on these expressions will be smaller than those based on the standard procedures for traits with a high level of dominance such as yield. Responses to five cycles of half-sib selection were predicted for maize by both expressions, considering that 100 progenies were evaluated and 10 S1 progenies were recombined, which corresponds to Ne = 10 for each cycle. The accumulated response to selection estimated with the new expression was about 47 and 28% smaller than that based on the standard expression for yield and plant height, respectively. Thus, the expression usually used overestimates the responses to selection, which is in agreement with reported results, because it does not take into account the effective population size that is generally small in recurrent selection programs

Alocação de linhagens de milho derivadas das populações BR-105 e BR-106 em grupos heteróticos

ABSTRACT: The heterotic groups are important in maize breeding programs because they allow the mostefficient use of the germoplasm. The objective of this research was to allocate maize lines to heterotic groupsfrom estimates of specific combining ability (SCA). Eight and ten S 3 lines derived from populations BR-105and BR-106, respectively, were crossed at interpopulation levels following a diallel system. Eighty single-crosses were obtained and evaluated in lattice designs across three environments. Recorded data consistedof grain yield (GY), plant height (PH) and ear height (EH). Estimates of general and specific combining ability(GCA) and (SGA) were computed according to Griffing’s method 4, model I, for all traits. Estimates of SCAwere used for principal coordinate analysis (PCO) and for cluster analysis, by using UPGMA (unweightedpair-group method with arithmetical averages) clustering algorithm, to assign the lines to heterotic groups. Forthe trait GY the lines were allocated to 4 heterotic groups, in which the lines from each population weresubdivided in 2 heterotic groups. For the traits PH and EH, the use of the estimates of SCA to assign lines toheterotic groups were not efficient. Cluster analysis and principal coordinate analysis were efficient for the traitGY in allocating maize lines to heterotic groups. Thus, since grain yield is the main trait for the breedingpurposes, with the allocation of the lines in 4 heterotic groups, the crosses will be more efficient, avoiding toobtain and evaluate unnecessary (hybrid) crosses.Key words: Zea mays, diallel analysis, combining ability

Genetic analysis of kernel oil content in tropical maize with design III and QTL mapping

Oil content and grain yield in maize are negatively correlated, and so far the development of high-oil high-yielding hybrids has not been accomplished. Then a fully understand of the inheritance of the kernel oil content is necessary to implement a breeding program to improve both traits simultaneously. Conventional and molecular marker analyses of the design III were carried out from a reference population developed from two tropical inbred lines divergent for kernel oil content. The results showed that additive variance was quite larger than the dominance variance, and the heritability coefficient was very high. Sixteen QTL were mapped, they were not evenly distributed along the chromosomes, and accounted for 30.91% of the genetic variance. The average level of dominance computed from both conventional and QTL analysis was partial dominance. The overall results indicated that the additive effects were more important than the dominance effects, the latter were not unidirectional and then heterosis could not be exploited in crosses. Most of the favorable alleles of the QTL were in the high-oil parental inbred, which could be transferred to other inbreds via marker-assisted backcross selection. Our results coupled with reported information indicated that the development of high-oil hybrids with acceptable yields could be accomplished by using marker-assisted selection involving oil content, grain yield and its components. Finally, to exploit the xenia effect to increase even more the oil content, these hybrids should be used in the Top Cross™ procedure.

Isoenzymatic variability in tropical maize populations under reciprocal recurrent selection

Maize (Zea mays L.) is one of the crops in which the genetic variability has been extensively studied at isoenzymatic loci. The genetic variability of the maize populations BR-105 and BR-106, and the synthetics IG-3 and IG-4, obtained after one cycle of a high-intensity reciprocal recurrent selection (RRS), was investigated at seven isoenzymatic loci. A total of twenty alleles were identified, and most of the private alleles were found in the BR-106 population. One cycle of reciprocal recurrent selection (RRS) caused reductions of 12% in the number of alleles in both populations. Changes in allele frequencies were also observed between populations and synthetics, mainly for the Est 2 locus. Populations presented similar values for the number of alleles per locus, percentage of polymorphic loci, and observed and expected heterozygosities. A decrease of the genetic variation values was observed for the synthetics as a consequence of genetic drift effects and reduction of the effective population sizes. The distribution of the genetic diversity within and between populations revealed that most of the diversity was maintained within them, i.e. BR-105 x BR-106 (GST = 3.5%) and IG-3 x IG-4 (GST = 4.0%). The genetic distances between populations and synthetics increased approximately 21%. An increase in the genetic divergence between the populations occurred without limiting new selection procedures.