BXD Recombinant Inbred Mice as a Model to Study Neurotoxicity

BXD recombinant inbred (RI) lines represent a genetic reference population derived from a cross between C57BL/6J mice (B6) and DBA/2J mice (D2), which through meiotic recombination events possesses recombinant chromosomes containing B6 or D2 haplotype segments. The quantitative trait loci (QTLs) are the locations of segregating genetic polymorphisms and are fundamental to understanding genetic diversity in human disease susceptibility and severity. QTL mapping represents the typical approach for identifying naturally occurring polymorphisms that influence complex phenotypes. In this process, genotypic values at markers of known genomic locations are associated with phenotypic values measured in a segregating population. Indeed, BXD RI strains provide a powerful tool to study neurotoxicity induced by different substances. In this review, we describe the use of BXD RI lines to understand the underlying mechanisms of neurotoxicity in response to ethanol and cocaine, as well as metals and pesticide exposures.

A Reevaluation of the Effect of Dietary Restriction on Different Recombinant Inbred (RI) Lines of Male and Female Mice

Dietary restriction (DR) was reported to either have no effect or reduced the lifespan of the majority of the 41-recombinant inbred (RI)-lines studied (Liao et al., 2010). In an appropriately power longevity study (n > 30 mice/group), we measured the lifespan of the four RI-lines (115-RI, 97-RI, 98-RI, and 107-RI) that were reported to have the greatest decrease in lifespan when fed 40% DR. DR increased the median lifespan of female and male 115-RI mice and female 97-RI and 107-RI mice. DR had little effect (less than 4%) on the median lifespan of female and male 98-RI mice and male 97-RI mice and reduced the lifespan of male 107-RI mice over 20%. While our study was unable to replicate the effect of DR on the lifespan of the RI-mice (except male 107-RI mice) reported by Liao et al. (2010), we found that the genotype of a mouse had a major impact on the effect of DR on lifespan, with the effect of DR ranging from a 50% increase to a 22% decrease. No correlation was observed between the changes in either body composition or glucose tolerance induced by DR and the changes observed in lifespan of the four RI-lines of male and female mice. These four RI-lines of mice give the research community a unique resource where investigators for the first time can study the anti-aging mechanism of DR by comparing mice in which DR increases lifespan to mice where DR has either no effect or reduces lifespan.

Performance of LS97-1610בSpencer’ soybean recombinant inbred line population segregating for resistance to Fusarium virguliforme

Clark, W. D., Reyes-Valdes, M. H., Bond, J. and Kantartzi, S. K. 2013. Performance of LS97-1610בSpencer’ soybean recombinant inbred line population segregating for resistance to Fusarium virguliforme . Can. J. Plant Sci. 93: 1179–1185. Sudden death syndrome (SDS) is a devastating disease in soybean which is caused by Fusarium virguliforme. Sudden death syndrome resistance is a quantitative trait; therefore, development of resistant varieties requires understanding of complex genetics and environmental effect. In this study, we aimed to characterize 94 F5:8 recombinant inbred (RI) lines derived from a cross between a resistant line for SDS, LS97-1610 and a susceptible cultivar, ‘Spencer’ and to identify sources of resistance. The RI lines were evaluated for their relative resistance (RR) to SDS along with the parents in four different environments; two years (2009 and 2010) and two locations (Carbondale and Valmeyer, IL). Analysis of variance revealed significant interactions between genotype and environment in each year of experimentation and in 2-yr combined data. Broad-sense heritability was estimated by ANOVA results, and it was moderate (61%). These results indicate that SDS resistance is partially unpredictable due to environmental influence. Ten RI were identified to have a RR with no significant differences from LS97-1610 and two of them had a RR lower than resistant parent. These stable RI across environments were used for further evaluation in breeding programs. Additionally, data obtained from field evaluation can be used in combination with molecular data to study the effect of quantitative trait loci (QTL) with the environment. The identification of common QTL across environments with consistent expression is the ultimate goal of every marker-assisted selection program.

System Approach to Understanding the Metabolic Diversity in Melon

Fruit quality is determined by numerous genetic factors that affect taste, aroma, ‎color, texture, nutritional value and shelf life. To unravel the genetic components ‎involved in the metabolic pathways behind these traits, the major goal of the project was to identify novel genes that are involved in, or that regulate, these pathways using correlation analysis between genotype, metabolite and gene expression data. The original and specific research objectives were: (1) Collection of replicated fruit from a population of 96 RI lines derived from parents distinguished by great diversity in fruit development and quality phenotypes, (2) Phenotypic and metabolic profiling of mature fruit from all 96 RI lines and their parents, (3) 454 pyrosequencing of cDNA representing mRNA of mature fruit from each line to facilitate gene expression analysis based on relative EST abundance, (4) Development of a database modeled after an existing database developed for tomato introgression lines (ILs) to facilitate online data analysis by members of this project and by researchers around the world. The main functions of the database will be to store and present metabolite and gene expression data so that correlations can be drawn between variation in target traits or metabolites across the RI population members and variation in gene expression to identify candidate genes which may impact phenotypic and chemical traits of interest, (5) Selection of RI lines for segregation and/or hybridization (crosses) analysis to ascertain whether or not genes associated with traits through gene expression/metabolite correlation analysis are indeed contributors to said traits. The overall research strategy was to utilize an available recombinant inbred population of melon (Cucumis melo L.) derived from phenotypically diverse parents and for which over 800 molecular markers have been mapped for the association of metabolic trait and gene expression QTLs. Transcriptomic data were obtained by high throughput sequencing using the Illumina platform instead of the originally planned 454 platform. The change was due to the fast advancement and proven advantages of the Illumina platform, as explained in the first annual scientific report. Metabolic data were collected using both targeted (sugars, organic acids, carotenoids) and non-targeted metabolomics analysis methodologies. Genes whose expression patterns were associated with variation of particular metabolites or fruit quality traits represent candidates for the molecular mechanisms that underlie them. Candidate genes that may encode enzymes catalyzingbiosynthetic steps in the production of volatile compounds of interest, downstream catabolic processes of aromatic amino acids and regulatory genes were selected and are in the process of functional analyses. Several of these are genes represent unanticipated effectors of compound accumulation that could not be identified using traditional approaches. According to the original plan, the Cucurbit Genomics Network (http://www.icugi.org/), developed through an earlier BARD project (IS-3333-02), was expanded to serve as a public portal for the extensive metabolomics and transcriptomic data resulting from the current project. Importantly, this database was also expanded to include genomic and metabolomic resources of all the cucurbit crops, including genomes of cucumber and watermelon, EST collections, genetic maps, metabolite data and additional information. In addition, the database provides tools enabling researchers to identify genes, the expression patterns of which correlate with traits of interest. The project has significantly expanded the existing EST resource for melon and provides new molecular tools for marker-assisted selection. This information will be opened to the public by the end of 2013, upon the first publication describing the transcriptomic and metabolomics resources developed through the project. In addition, well-characterized RI lines are available to enable targeted breeding for genes of interest. Segregation of the RI lines for specific metabolites of interest has been shown, demonstrating the utility in these lines and our new molecular and metabolic data as a basis for selection targeting specific flavor, quality, nutritional and/or defensive compounds. To summarize, all the specific goals of the project have been achieved and in many cases exceeded. Large scale trascriptomic and metabolomic resources have been developed for melon and will soon become available to the community. The usefulness of these has been validated. A number of novel genes involved in fruit ripening have been selected and are currently being functionally analyzed. We thus fully addressed our obligations to the project. In our view, however, the potential value of the project outcomes as ultimately manifested may be far greater than originally anticipated. The resources developed and expanded under this project, and the tools created for using them will enable us, and others, to continue to employ resulting data and discoveries in future studies with benefits both in basic and applied agricultural - scientific research.

IDENTIFICATION OF A MAJOR QUANTITATIVE TRAIT LOCUS CONFERRING RICE BLAST RESISTANCE USING RECOMBINANT INBRED LINES

Blast disease caused by Pyricularia oryzae is one of the limiting factors for rice production world wide. The use of resistant varieties for managing blast disease is considered as the most eco-friendly approaches. However, their resistances may be broken down within a few years due to the appearance of new virulent blast races in the field. The objective of the present study was to identify the quantitative trait locus (QTL) conferring resistance to blast disease using 126 recombinant inbred (RI) lines originated from a crossing of a durably resistant upland rice genotype (Laka) and a highly susceptible rice accession cultivar (Kencana Bali). The RI population was developed through a single seed descent method from 1997 to 2004. Resistance of the RI lines was evaluated for blast in an endemic area of Sukabumi, West Java, in 2005. Disease intensity of the blast was examined following the standard evaluation system developed by the International Rice Research Institute (IRRI). At the same year the RI lines were analyzed with 134 DNA markers. Results of the study showed that one major QTL was found to be associated with blast resistance, and this QTL was located near RM2136 marker on the long arm of chromosome 11. This QTL explained 87% of the phenotypic variation with 37% additive effect. The map position of this QTL differed from that of a partial resistant gene, Pi34, identified previously on chromosome 11 in the Japanese durably resistant variety, Chubu 32. The QTL, however, was almost at the same position as that of the multiple allele-resistant gene, Pik. Therefore, an allelic test should be conducted to clarify the allelic relationship between QTL identified in this study and the Pik. The RI lines are the permanent segregating population that could be very useful for analysing phenotypic variations of important agronomic traits possibly owned by the RI lines. The major QTL identified in this study could be used as a genetic resource in improvement of rice varieties for blast resistance in Indonesia

IDENTIFICATION OF A MAJOR QUANTITATIVE TRAIT LOCUS CONFERRING RICE BLAST RESISTANCE USING RECOMBINANT INBRED LINES

Blast disease caused by Pyricularia oryzae is one of the limiting<br />factors for rice production world wide. The use of resistant<br />varieties for managing blast disease is considered as the most<br />eco-friendly approaches. However, their resistances may be<br />broken down within a few years due to the appearance of new<br />virulent blast races in the field. The objective of the present<br />study was to identify the quantitative trait locus (QTL) conferring<br />resistance to blast disease using 126 recombinant inbred<br />(RI) lines originated from a crossing of a durably resistant upland<br />rice genotype (Laka) and a highly susceptible rice accession<br />cultivar (Kencana Bali). The RI population was developed<br />through a single seed descent method from 1997 to 2004.<br />Resistance of the RI lines was evaluated for blast in an endemic<br />area of Sukabumi, West Java, in 2005. Disease intensity of the<br />blast was examined following the standard evaluation system<br />developed by the International Rice Research Institute (IRRI).<br />At the same year the RI lines were analyzed with 134 DNA<br />markers. Results of the study showed that one major QTL was<br />found to be associated with blast resistance, and this QTL was<br />located near RM2136 marker on the long arm of chromosome<br />11. This QTL explained 87% of the phenotypic variation with<br />37% additive effect. The map position of this QTL differed<br />from that of a partial resistant gene, Pi34, identified previously<br />on chromosome 11 in the Japanese durably resistant variety,<br />Chubu 32. The QTL, however, was almost at the same position<br />as that of the multiple allele-resistant gene, Pik. Therefore, an<br />allelic test should be conducted to clarify the allelic relationship<br />between QTL identified in this study and the Pik. The RI lines<br />are the permanent segregating population that could be very<br />useful for analysing phenotypic variations of important agronomic<br />traits possibly owned by the RI lines. The major QTL<br />identified in this study could be used as a genetic resource in<br />improvement of rice varieties for blast resistance in Indonesia

Maize Leaf Epiphytic Bacteria Diversity Patterns Are Genetically Correlated with Resistance to Fungal Pathogen Infection

Plant leaves host a specific set of microbial epiphytes. Plant genetic and solar UV-B radiation effects on the diversity of the phyllosphere were examined by measuring epiphytic bacterial ribosomal DNA diversity in a maize recombinant inbred (RI) mapping population. Several chromosomal quantitative trait loci (QTL) with significant effects on bacterial diversity were identified, some of which had effects only in the presence of UV-B radiation and others that had effects both with and without UV-B. Candidate genes with allele-specific effects were mapped to the bacterial diversity chromosomal regions. A glutamate decarboxylase candidate gene was located at a UV-B–specific chromosomal locus, and in a comparison between two RI lines with contrasting bacterial diversity phenotypes, high bacterial diversity was associated with high levels of glutamate decarboxylase enzyme activity, a component of the gamma-aminobutyric acid (GABA) pathway. The bacterial diversity loci exhibited a significant overlap with loci connected with Southern leaf blight (SLB) susceptibility in the field. A SLB-resistant inbred genotype had less beta bacterial diversity, and antibiotic treatment of inbreds increased this diversity. These results suggest that the GABA pathway is genetically associated with phyllosphere bacterial diversity. Furthermore, the colocalization of QTL between low bacterial diversity and fungal blight–resistance and the increase in beta diversity in antibiotic-treated leaves suggest that occupation of leaf habitats by a particular set of suppressive bacteria may restrict phyllosphere bacterial variability and increase resistance to fungal infection.

Biomass Distribution in Kalanchoe blossfeldiana Transformed with rol-genes of Agrobacterium rhizogenes

Kalanchoe blossfeldiana transformed with Agrobacterium rhizogenes exhibited marked alterations in morphology and biomass distribution. Plants termed root-inducing (Ri) lines were regenerated from hairy roots produced by inoculating leaf explants with Agrobacterium rhizogenes wild-type strain ATCC15834. Six Ri lines were characterized in a greenhouse trial and all Ri lines had reduced dry weights of main shoot, lateral shoots, leaves, and flowers compared with control plants. The reduction in dry weights of these organs correlated with reduced plant height, shoot length, leaf area, and number of flowers per plant. Furthermore, an altered distribution of dry matter was evident in the Ri plants, where the greater part of dry matter was allocated into leaves and secondly into flowers, whereas the majority of dry matter in control plants was allocated into flowers and secondly into leaves. Furthermore, a higher percentage of dry matter was allocated into the main shoot of the Ri lines in comparison with that of control plants. Increased dry matter in leaves and in the main shoot in the Ri lines appeared to be at the expense of dry matter allocated into flowers. Moreover, an increased number of vegetative lateral shoots was recorded in the Ri lines, whereas the number of reproductive lateral shoots was decreased. Possible mechanisms behind the altered resource distribution are discussed.

Quantitative Trait Loci Conditioning Resistance to Phaeosphaeria Leaf Spot of Maize Caused by Phaeosphaeria maydis

Phaeosphaeria leaf spot (PLS) is a potentially important disease of maize (Zea mays) that has appeared in winter breeding nurseries in southern Florida. Inbred lines related to B73 are particularly susceptible to Phaeosphaeria leaf spot, whereas inbreds related to Mo17 are highly resistant. A previous study of the inheritance of resistance to Phaeosphaeria leaf spot in the cross B73 × Mo17 found that resistance is highly heritable and controlled by mostly additive gene action at three or four loci. In this study, we used 158 recombinant inbred (RI) lines derived from the cross B73 × Mo17 to map quantitative trait loci (QTL) governing resistance. The RI lines along with the parent inbred lines and the F1 were evaluated for PLS resistance in replicated trials over two winter growing seasons in southern Florida. Using the composite interval mapping (CIM) function of PLABQTL software, five QTL on four different chromosomes were found to control PLS resistance in Mo17. In addition, the × additive interaction between two of these QTL was found to be significant. Our results are in close agreement with the previous study, where generation mean analysis was used to study the inheritance of resistance to PLS.