Analysis of genomic and functional RFLP derived markers associated with sucrose content, fiber and yield QTLs in a sugarcane (Saccharum spp.) commercial cross

Expressed sequence tags derived markers have a great potential to be used in functional map construction and QTL tagging. In the present work, sugarcane genomic probes and expressed sequence tags having homology to genes, mostly involved in carbohydrate metabolism were used in RFLP assays to identify putative QTLs as well as their epistatic interactions for fiber content, cane yield, pol and tones of sugar per hectare, at two crop cycles in a progeny derived from a bi-parental cross of sugarcane elite materials. A hundred and twenty marker trait associations were found, of which 26 at both crop cycle and 32 only at first ratoon cane. A sucrose synthase derived marker was associated with a putative QTL having a high negative effect on cane yield and also with a QTL having a positive effect on Pol at both crop cycles. Fifty digenic epistatic marker interactions were identified for the four traits evaluated. Of these, only two were observed at both crop cycles.

Genetic variability among sugarcane genotypes based on polymorphisms in sucrose metabolism and drought tolerance genes

Target region amplification polymorphism (TRAP) markers were used to estimate the genetic similarity (GS) among 53 sugarcane varieties and five species of the Saccharum complex. Seven fixed primers designed from candidate genes involved in sucrose metabolism and three from those involved in drought response metabolism were used in combination with three arbitrary primers. The clustering of the genotypes for sucrose metabolism and drought response were similar, but the GS based on Jaccard`s coefficient changed. The GS based on polymorphism in sucrose genes estimated in a set of 46 Brazilian varieties, all of which belong to the three Brazilian breeding programs, ranged from 0.52 to 0.9, and that based on drought data ranged from 0.44 to 0.95. The results suggest that genetic variability in the evaluated genes was lower in the sucrose metabolism genes than in the drought response metabolism ones.

Evaluation of algorithms used to order markers on genetic maps

When building genetic maps, it is necessary to choose from several marker ordering algorithms and criteria, and the choice is not always simple. In this study, we evaluate the efficiency of algorithms try (TRY), seriation (SER), rapid chain delineation (RCD), recombination counting and ordering (RECORD) and unidirectional growth (UG), as well as the criteria PARF (product of adjacent recombination fractions), SARF (sum of adjacent recombination fractions), SALOD (sum of adjacent LOD scores) and LHMC (likelihood through hidden Markov chains), used with the RIPPLE algorithm for error verification, in the construction of genetic linkage maps. A linkage map of a hypothetical diploid and monoecious plant species was simulated containing one linkage group and 21 markers with fixed distance of 3 cM between them. In all, 700 F(2) populations were randomly simulated with and 400 individuals with different combinations of dominant and co-dominant markers, as well as 10 and 20% of missing data. The simulations showed that, in the presence of co-dominant markers only, any combination of algorithm and criteria may be used, even for a reduced population size. In the case of a smaller proportion of dominant markers, any of the algorithms and criteria (except SALOD) investigated may be used. In the presence of high proportions of dominant markers and smaller samples (around 100), the probability of repulsion linkage increases between them and, in this case, use of the algorithms TRY and SER associated to RIPPLE with criterion LHMC would provide better results. Heredity (2009) 103, 494-502; doi:10.1038/hdy.2009.96; published online 29 July 2009

Characterization of new polymorphic functional markers for sugarcane

Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)[02/01167-1]

An integrated molecular map of yellow passion fruit based on simultaneous maximum-likelihood estimation of linkage and linkage phases

The development of genetic maps for auto-incompatible species, such as the yellow passion fruit (Passiflora edulis Sims f.flavicarpa Deg.) is restricted due to the unfeasibility of obtaining traditional mapping populations based on inbred lines. For this reason, yellow passion fruit linkage maps were generally constructed using a strategy known as two-way pseudo-testeross, based on monoparental dominant markers segregating in a 1:1 fashion. Due to the lack of information from these markers in one of the parents, two individual (parental) maps were obtained. However, integration of these maps is essential, and biparental markers can be used for such an operation. The objective of our study was to construct an integrated molecular map for a full-sib population of yellow passion fruit combining different loci configuration generated from amplified fragment length polymorphisms (AFLPs) and microsatellite markers and using a novel approach based on simultaneous maximum-likelihood estimation of linkage and linkage phases, specially designed for outcrossing species. Of the total number of loci, approximate to 76%, 21%, 0.7%, and 2.3% did segregate in 1:1, 3:1, 1:2:1, and 1:1:1:1 ratios, respectively. Ten linkage groups (LGs) were established with a logarithm of the odds (LOD) score >= 5.0 assuming a recombination fraction : <= 0.35. On average, 24 markers were assigned per LG, representing a total map length of 1687 cM, with a marker density of 6.9 cM. No markers were placed as accessories on the map as was done with previously constructed individual maps.

Influence of egg pre-storage heating period and storage length on incubation results

This experiment aimed at evaluating the influence of different heating times of settable eggs of Cobb 500 (R) broiler breeders before submitting them to different storage periods on egg weight loss, embryo mortality, and hatchability. A total number of 1,980 eggs were distributed in a completely randomized experimental design with a 3 x 3 factorial arrangement, comprising nine treatments with 22 replicates of 10 eggs each. The following factors were analyzed: pre-storage heating periods (0, 6, 12 hours at 36.92 degrees C) and storage periods (4, 9, 14 days at 12.06 degrees C). After storage, eggs were incubated under usual conditions, and were transferred to the hatcher at 442 hours of incubation. Eggs were weighed before heating, incubation, and transference to determine weight loss. Partial hatchability was determined at 480 hours, and total hatchability at 498 hours of incubation. Embryo mortality was determined in non-hatched eggs. It was concluded that heating eggs for six hour before storage improves incubation results as it decreases incubation length and late embryo mortality, therefore its use can be indicated in commercial operations. Storing eggs for 14 days and pre-heating for 14 days and pre-heating for 12 hours severely impair incubation results, and therefore are not recommended.

Influence of egg pre-storage heating period and storage length on the digestive tract of newly-hatched broiler chicks

An experiment was carried out to evaluate the effect of different heating times of settable eggs of Cobb 500 (R) broiler breeders before submitting them to different storage periods on body weight, digestive tract organ weights, and intestinal mucosa morphology of newly-hatched chicks. Settable eggs were distributed in a completely randomized experimental design with a 3 x 3 factorial arrangement: pre-storage heating periods (0, 6, 12 hours at 36.92 degrees C) and storage periods (4, 9, 14 days at 12.06 degrees C). Body weight and relative weights of the yolk sac, heart, liver, proventriculus+gizzard, and intestinal segments were measured in chicks hatching at 480 and 498 hours of incubation. Villi height, width and perimeter, and crypt depth < im) were measured in duodenal histological sections. It was concluded that pre-storage healing for six hours of eggs stored for four or nine days increases small intestine weight of newly-hatched chicks, but does not influence the morphology of the duodenal mucosa. Pre-storage heating for 12 hours negatively influences body weight and duodenal mucosa development, and therefore this practice is not recommended. Storage length does not have consistent effect on body weight and development of the gastrointestinal tract.

Inheritance of growth habit detected by genetic linkage analysis using microsatellites in the common bean (Phaseolus vulgaris L.)

The genetic linkage map for the common bean (Phaseolus vulgaris L.) is a valuable tool for breeding programs. Breeders provide new cultivars that meet the requirements of farmers and consumers, such as seed color, seed size, maturity, and growth habit. A genetic study was conducted to examine the genetics behind certain qualitative traits. Growth habit is usually described as a recessive trait inherited by a single gene, and there is no consensus about the position of the locus. The aim of this study was to develop a new genetic linkage map using genic and genomic microsatellite markers and three morphological traits: growth habit, flower color, and pod tip shape. A mapping population consisting of 380 recombinant F10 lines was generated from IAC-UNA x CAL143. A total of 871 microsatellites were screened for polymorphisms among the parents, and a linkage map was obtained with 198 mapped microsatellites. The total map length was 1865.9 cM, and the average distance between markers was 9.4 cM. Flower color and pod tip shape were mapped and segregated at Mendelian ratios, as expected. The segregation ratio and linkage data analyses indicated that the determinacy growth habit was inherited as two independent and dominant genes, and a genetic model is proposed for this trait.

A new polymorphism in the Growth and Differentiation Factor 9 (GDF9) gene is associated with increased ovulation rate and prolificacy in homozygous sheep

P>Brazilian Santa Ines (SI) sheep are very well-adapted to the tropical conditions of Brazil and are an important source of animal protein. A high rate of twin births was reported in some SI flocks. Growth and Differentiation Factor 9 (GDF9) and Bone Morphogenetic Protein 15 (BMP15) are the first two genes expressed by the oocyte to be associated with an increased ovulation rate in sheep. All GDF9 and BMP15 variants characterized, until now, present the same phenotype: the heterozygote ewes have an increased ovulation rate and the mutated homozygotes are sterile. In this study, we have found a new allele of GDF9, named FecGE (Embrapa), which leads to a substitution of a phenylalanine with a cysteine in a conservative position of the mature peptide. Homozygote ewes presenting the FecGE allele have shown an increase in their ovulation rate (82%) and prolificacy (58%). This new phenotype can be very useful in better understanding the genetic control of follicular development; the mechanisms involved in the control of ovulation rate in mammals; and for the improvement of sheep production.

A new set of microsatellite markers for the genetic characterization of Ceratocystis fimbriata, an economically important plant pathogen

Ceratocystis fimbriata is a fungal pathogen which attacks several economically important plants, but occurs in host-associated, morphologically indistinguishable forms. In Brazil, this fungus seriously attacks mango trees (Mangifera indica), causing severe loss of yield. This work aimed to develop and characterize a novel set of microsatellite markers for this important pathogen, providing researchers with new molecular tools for the characterization of isolates. Twenty polymorphic primer pairs were designed from a microsatellite-enriched library. We tested the usefulness of these markers through genotyping thirteen isolates of the fungus. On average, 6.65 alleles per locus were detected, revealing the ability of this set of markers to characterize C. fimbriata isolates associated to mango and to other plant species.

Outcrossing rate in sweet passion fruit based on molecular markers

P>Yellow and sweet passion fruit are insect-pollinated species native to the tropics. Fruits are used commercially for human consumption worldwide. The yellow passion fruit is an outcrossing species with self-incompatible flowers. However, the reproductive system of the sweet passion fruit (Passiflora alata) has not been well elucidated. The objective of this work was to characterize aspects of the mating system in the sweet passion fruit using random amplified polymorphic DNA (RAPD) and microsatellite markers, particularly the rate of outcrossing in P. alata progenies. A multilocus outcrossing rate of t(m) = 0.994 was determined from RAPD and t(m) = 0.940 from microsatellites, supporting P. alata as an outcrossing species. The fixation indices of the maternal generation (F(m)) were -0.200 and 0.071 with RAPD and microsatellite loci, respectively, indicating the absence of inbreeding in the maternal generation. The paternity correlation (r(p)) varied from -0.008 with RAPD markers to 0.208 with microsatellite markers, suggesting a low probability of finding full sibs within the progenies. The results demonstrated that all progenies assessed in this study were derived from outcrossing.

Development and characterization of microsatellite markers for turmeric (Curcuma longa)

P>Curcuma longa L. is a sterile, triploid, vegetatively-propagated crop cultivated mainly in Southeast Asia. When dried rhizomes are ground, the resulting yellow powder is used by the food industry as a natural food dye. Moreover, many pharmacological compounds have broadened the commercial application of the crop. However, conventional breeding is difficult and hence, improvement has been limited to germplasm selection. To better utilize the germplasm collections and facilitate genotype selection, a total of 17 polymorphic microsatellite loci were developed using a CT/GT/CTT enriched genomic library. All microsatellites resulted in amplified PCR products, showing a banding pattern of 2-11 polymorphic bands per locus, enabling genotype discrimination. These results can be used in further studies aimed at characterizing C. longa genetic resource collections and also to improve breeding strategies.

Development and characterization of microsatellite markers from Guarea guidonia (Meliaceae), a tree species from different habitats within the Brazilian Atlantic forest

The tree species Guarea guidonea (Meliaceae) belongs to a predominantly tropical family, being largely found in natural or anthropic forest fragments within the Brazilian Atlantic Forest. Aiming to develop future studies on the genetic structure of plant species from forests fragments, eleven microsatellite markers were developed for Guarea guidonia, based on the analysis of 45 individuals from natural populations of three different fragments within the forest-anthropic edge, interior fragment and natural edge. Only eight loci showed to be polymorphic and the number of alleles ranged from two to four (mean of 2.50). All populations showed almost the same level of genetic diversity (mean H(e) = 0.3775). These loci will be useful for population genetics studies on Guarea guidonea, providing information for the conservation and management of this species.

Development and characterization of microsatellite markers for castor (Ricinus communis L.), an important oleaginous species for biodiesel production

Castor (Ricinus communis L.) is an important oleaginous plant from both economic and social points of view. The seeds contain an oil with excellent properties for industrial uses. This paper presents the main results of a study aiming to develop microsatellite markers for castor. Twelve new polymorphic microsatellite markers were isolated and characterized in 38 genotypes accessions from the castor germplasm of the Brazilian Agricultural Research Company (EMBRAPA). Knowledge on the genetic diversity of castor can be used to gain a better understanding on genetic diversity conservation, and germplasm management, guiding breeding programs and conservation strategies.

Development and characterization of microsatellite markers for Psychotria tenuinervis (Rubiaceae), a shrub species from the Atlantic forest, and primers transferability from Coffea

The shrub species Psychotria tenuinervis (Rubiaceae) is native to the Brazilian Atlantic forest and is largely found within natural and disturbed forest fragments. Aiming to develop studies on population genetic structure of forest fragment species, eigth microsatellite markers were developed for P. tenuinervis. Also, 15 loci already developed for Coffea (Rubiaceae) were tested for transferability to this species. We utilized 45 individuals from natural populations of three different fragments-anthropic edge, interior fragment and natural edge, within the Brazilian Atlantic forest. The average number of alleles per locus was 2.5 (two-four alleles/locus). These loci will be useful for future population genetic studies aiming to the conservation and management of this species.