An F2 Barley Population as a Tool for Teaching Mendelian Genetics

Plant genetics is more quantitative and analytical than other areas of plant biology. The mathematical descriptions of Mendelian inheritance and population genetics are sometimes discouraging, and students often have serious misconceptions. Innovative strategies in expositive classes can clearly encourage student's motivation and participation, but laboratories and practical classes are generally the students' most favorite academic activities. Genetic practices are usually employed with the aim to teach experimental methods (as PCR, DNA extraction,…). However, the design of lab practices focused on the learning of abstract concepts such as genetic interaction, quantitative inheritance, genetic linkage, genetic recombination, gene mapping, or molecular markers, is a complex task that requires a suitable segregant plant material. The most optimal population for pedagogical purposes is an F2 population, which results in extremely useful not only to explain different key concepts of genetics (as dominance, epistasis, and linkage) but also to introduce to the student's additional curricular tools, particularly, concerning statistical analysis. Among various model organisms available, barley possesses several unique features for demonstrating genetic principles. Therefore, we have generated a barley F2 population from the parental lines of the Oregon Wolfe Barley collection (https://barleyworld.org/owb). The objective of this work is to present this F2 population, composed of more than 300 individuals, as a model to teach Mendelian genetics in a medium-high level Genetics course. We provide an exhaustive phenotypic and genotypic description of this plant material, including qualitative and quantitative traits as well as a set of molecular markers. Examples of genetic interaction (epistasis) and linkage analysis are explored and discussed. The description of the specific methodologies and practical exercises carried out in our Genetics courses can be helpful for transferring our fruitful experience to anyone interested in implementing this educational resource in his/her teaching.

Phytophthora capsici Leonian causes significant yield losses in commercial squash (Cucurbita pepo) production worldwide. The deployment of resistant cultivars can complement integrated management practices for P. capsici, but resistant cultivars are currently unavailable for growers. Moderate resistance to Phytophthora crown rot in a selection of accession PI 181761 (C. pepo) (designated line #181761-36P) is controlled by three dominant genes (R4, R5 and R6). Introgression of these loci into elite germplasm through marker-assisted selection (MAS) can accelerate the release of new C. pepo cultivars resistant to crown rot, but these tools are currently unavailable. Here we describe the identification of a quantitative trait locus (QTL), molecular markers and candidate genes associated with crown rot resistance in #181761-36P. Five hundred and twenty-three SNP markers were genotyped in an F2 (n = 83) population derived from a cross between #181761-36P (R) and Table Queen (S) using targeted genotyping by sequencing. A linkage map (2068.96 cM) consisting of twenty-one linkage groups and an average density of 8.1 markers/cM was developed for the F2 population. The F2:3 families were phenotyped in the greenhouse with a virulent strain of P. capsica, using the spore-spray method. A single QTL (QtlPC-C13) was consistently detected on LG 13 (chromosome 13) across three experiments and explained 17.92–21.47% of phenotypic variation observed in the population. Nine candidate disease resistance gene homologs were found within the confidence interval of QtlPC-C13. Single nucleotide polymorphism (SNP) markers within these genes were converted into Kompetitive Allele Specific PCR (KASP) assays and tested for association with resistance in the F2 population. One SNP marker (C002686) was significantly associated with resistance to crown rot in the F2 population (p < 0.05). This marker is a potential target for MAS for crown rot resistance in C. pepo.

BackgroundThe availability of a large expressed sequence tags (EST) resource and recent advances in high-throughput genotyping technology have made it possible to develop highly multiplexed SNP arrays for multi-objective genetic applications, including the construction of meiotic maps. Such approaches are particularly useful in species with a large genome size, precluding the use of whole-genome shotgun assembly with current technologies.ResultsIn this study, a 12 k-SNP genotyping array was developed for maritime pine from an extensive EST resource assembled into a unigene set. The offspring of three-generation outbred and inbred mapping pedigrees were then genotyped. The inbred pedigree consisted of a classical F2 population resulting from the selfing of a single inter-provenance (Landes x Corsica) hybrid tree, whereas the outbred pedigree (G2) resulted from a controlled cross of two intra-provenance (Landes x Landes) hybrid trees. This resulted in the generation of three linkage maps based on SNP markers: one from the parental genotype of the F2 population (1,131 markers in 1,708 centimorgan (cM)), and one for each parent of the G2 population (1,015 and 1,110 markers in 1,447 and 1,425 cM for the female and male parents, respectively). A comparison of segregation patterns in the progeny obtained from the two types of mating (inbreeding and outbreeding) led to the identification of a chromosomal region carrying an embryo viability locus with a semi-lethal allele. Following selfing and segregation, zygote mortality resulted in a deficit of Corsican homozygous genotypes in the F2 population. This dataset was also used to study the extent and distribution of meiotic recombination along the length of the chromosomes and the effect of sex and/or genetic background on recombination. The genetic background of trees in which meiotic recombination occurred was found to have a significant effect on the frequency of recombination. Furthermore, only a small proportion of the recombination hot- and cold-spots were common to all three genotypes, suggesting that the spatial pattern of recombination was genetically variable.ConclusionThis study led to the development of classical genomic tools for this ecologically and economically important species. It also identified a chromosomal region bearing a semi-lethal recessive allele and demonstrated the genetic variability of recombination rate over the genome.

New Technologies, Tools and Approaches for Improving Crop Breeding

BOOK REVIEWS Human Gene Mutation. By David N. Cooper and Michael Krawczak. Oxford: BIOS Scientific Publishers, 1993. Pp. 402. $99.00. Extensive libraries of monogenic mutations, detailed linkage maps and the domestication of certain prokaryotic and eukaryotic species with desirable technical advantages characterized the model species of transmission geneticists. The mouse was the model species for mammals but not in the same class as the other species. Homo sapiens was never viewed as a member of the inner circle. And then came molecular genetics! The extraordinary progress in molecular genetics, initially involving the model species, over the past fifteen years has transformed genetics and probably founded a new subdiscipline of biochemistry. Sophisticated gene manipulations are now commonplace and accomplished by kits, equipment, and technicians. Yet the raw materials of the transmission geneticists, genie mutations, remain the same as those of the transmission geneticists. Only one mammalian species has the largest number of identified monogenically determined mutations and the most detailed linkage maps: Homo sapiens. Molecular geneticists have been quick to tap federal financial resources (the human genome project) and the latest technical breakthroughs (identified by an arcane series of letters or acronyms) to publish their genie sequences in a small library of established and newly established journals. The need for a comprehensive and integrated overview has long been obvious. Cooper and Krawczak have written a welcome book which provides an overview of progress in the molecular genetics of human gene mutations. "The aim of the volume has been to put together under one cover an empirically based mechanistic analysis of the nature of human gene mutation causing genetic disease, an assessment of the consequences of mutation both at the level of gene expression and for the complex relationships between genotype and phenotype, an overview of the currently available methods for mutation detection and analysis , and a reference source to current diagnostic possibilities in a fast-moving field." Each of the thirteen chapters has comprehensive contemporary references . The information in numerous tables and four appendices are not likely to be found elsewhere. The line drawings and figures are designed to illuminate the text. The book could provide the definitive text for an advanced course in modern human genetics—after exposure to a strong introductory course in genetics and Permission to reprint a book review printed in this section may be obtained only from the author. Perspectives in Biology and Medicine, 37, 4 ¦ Summer 1994 609 molecular genetics. At the same time, it is an excellent resource for specific examples to illustrate concepts for a course in transmission and molecular genetics . The price is likely to discourage purchase for the personal library of teachers and graduate students interested or involved in human genetics, particularly at the molecular level, but should not discourage purchase for a departmental or institutional library. E. D. Garber Department of Ecology and Evolution University of Chicago Emerging Viruses. Edited by Stephen Morse. Oxford: Oxford Univ. Press, 1993. Pp. 317. $39.95. Man has been fascinated by epidemics since biblical times. "For now I will stretch out my hand that I may smite thee and thy people with pestilence . . ." (Exodus 9). Along with this fascination have come attempts to anticipate these epidemics and to prevent them. Stephen Morse's volume on Emerging Viruses is based on a conference that deals with these concerns. Some of the original speakers at the conference, experts in varied fields, describe virus epidemics, virus detection methods, the evolution of viruses, and recommendations regarding prevention and control of emerging viruses. These topics are especially timely because of the present HIV-I threat. It comes as a surprise to realize that this topic is actually a unique focus for a book and, as Lederberg notes in his chapter, "the present examination is virtually without precedent." Amidst the plethora of books that deal with over-worked topics related to infectious disease, this insightful volume is certainly welcome. It is readable as well as scholarly. The stage for the book is well-set by Morse's perceptive chapter "Examining the origins of emerging viruses." As noted by Morse, bona fide newly emerging viruses are generally new variants of an established virus recently introduced from another species or another...

The soybean aphid (Aphis glycines Matsumura) is a major pest on soybean [Glycine max (L.) Merr.] in North America. Aphid resistance has been found on plant introduction (PI) 567537, but its genetic characterization is unknown. The objectives of this study were to identify the resistance genes in PI 567537 using molecular markers and validate them in a different genetic background. A mapping population of 86 F4 lines from a cross between PI 567537 and a susceptible parent E00003 was investigated for aphid resistance in both greenhouse and field trials. A genomic region associated with the aphid resistance in PI 567537 was revealed on chromosome 16 (linkage group J) with molecular markers. This locus was coincidently located in the same region as Rag3 and explained most of the phenotypic variation, ranging from 87.4 % in the greenhouse trial to 78.9 % in the field trial. This resistance gene was further confirmed in an F2 population derived from a cross of PI 567537 × Skylla. The segregation of the F2 population indicated that the aphid resistance in PI 567537 was most likely controlled by a single dominant gene, which was the one we mapped in the F4-derived population. This gene was designated Rag3b since it is located in the same region as Rag3. The mapping of the aphid resistance gene in PI 567537 could be useful in marker-assisted selection when employing PI 567537 as an aphid resistance source.

Construction of SNP genetic maps based on targeted next-generation sequencing and QTL mapping of vital agronomic traits in faba bean (Vicia faba L.)

Rice grain shape, heading date, plant height and panicle length are the most important factors affecting rice grain quality, yield and adaption area. To detect QTLs controlling grain length (GL), grain width (GW), heading date (HD), plant height (PH), and panicle length (PL), we evaluated two hundred and eighty six F2 individuals derived from the cross between Taiken 2 (TK2) and Taichung Sen 10 (TCS10). Through genotyping all the F2 population, we construct a linkage map with 147 molecular markers including 133 InDel and 14 SSR markers covering all 12 chromosomes in this study. The phenotypic ranges of grain trait in F2 population were between two parents, but the other traits showed transgressive segregation. The correlation between GL and GW are only 0.09. The result of QTL mapping shows that there are 10 and 8 QTLs controlling GL and GW. The PVE of QTLs controlling GL on chromosome 1, 3, 7 are all above 10 %, and GL would increase 0.18 – 0.22 mm by carrying TCS 10 allele. The PVE of QTLs controlling GW on chromosome 2 and 5 are 12% and 44.8%, and GW would increase 0.08 mm and 0.17 mm by carrying TK2 allele. A total 3 QTLs for HD, 5 QTLs for PH and 4 QTLs for PL. The PVE of QTLs on chromosome 3 and 10 controlling HD are around 26%. While these two QTLs carrying TK2 allele, the HD would earlier 6.7 days and delay 6.3 day. The QTL controlling PH on chromosome 1 is the major QTL explained 52.5% of the total variation of PH, and it would increase 14.9 cm with TK2 allele on it. The QTL of PL on chromosome 8 has the largest PVE and would increase 1.5 cm while carrying TK2 allele. Compare all these result with previous study, the QTL of GW on chromosome 5 might be the GW5 gene, and the QTL controlling HD on chromosome 10 would be Ehd1. The QTL at the long arm on chromosome 1 might be the same as sd-1. All these assumption need to be confirm by functional markers or sequence alignment. All these information would be useful to the rice breeder or have practical used through MAS in rice.

Two related segregating populations of Theobroma cacao L. were analysed for their resistance to Phytophthora palmivora. The first F1 population was obtained by crossing two susceptible cacao clones of Catongo (a highly homozygous genotype) and Pound 12(a highly heterozygous genotype) and the second population was obtained by backcrossing a single F1 tree with Catongo. The genetic maps obtained for each population were compared. The F1 map includes 162 loci and the backcross has 140 loci. The two maps, F1 and BC1, exhibit high co-linear loci organization covering respectively, 772 and 944 cM.Phytophthora resistance was assessed by measuring the size increase of a lesion at five (DL5)and ten days (DL10) after pod inoculation. Six different QTL were detected in the F1 and BC1 populations. One QTL was found in both populations, and appeared to be a major component of disease resistance, and explaining nearly 48% of the phenotypic variance in the F1 population. The absence of some yield QTL detection in the BC1 in comparison with the F1 population is due to the lack of transmission of the favouring alleles for these QTL from the single F1 tree used for the backcross. The phenotypic variance explained by the action of the quantitative trait alleles indicated that genetic factors of both major and minor effects were involved in the control of the character studied. QTL conferring increased resistance to Phytophthorawere identified in both susceptible parents, suggesting the presence of transgressive traits and the possibility of selection in cacao. Pleiotropic and epistatic effects for the QTL were also detected. Finally, the use of marker assisted selection (MAS) in cacao breeding programs is discussed.

The development of a complete linkage map, including both classical (visible) and molecular markers, is important to understand the genetic relationships among different traits in common bean (Phaseolus vulgaris L.). The objective of this study was to integrate classical marker genes into previously constructed molecular linkage maps in common bean. Bulked segregant analysis was used to identify 10 random amplified polymorphic DNA (RAPD) markers linked to genes for five classical marker traits: dark green savoy leaf (dgs), blue flower (blu), silvery [Latin: argentum] green pod (arg), yellow wax pod (y) and flat pod (a spontaneous mutation from round to flat pod in `Hialeah' snap bean). The genes for dark green savoy leaf (dgs) and blue flower (blu) were located in a previously constructed molecular linkage map. These results indicate that classical marker genes and molecular markers can be integrated to form a more complete and informative genetic linkage map. Most of the RAPD markers were not polymorphic in the two mapping populations used, and molecular markers from those mapping populations were not polymorphic in the F2 populations used to develop the RAPD markers. Alternative genetic hypotheses for the pod shape mutation in `Hialeah' are discussed, and the experimental difficulties of pod shape classification are described.

Seed coat color is an important agronomic trait in sesame, as it is associated with seed biochemical properties, antioxidant content and activity and even disease resistance of sesame. Here, using a high-density linkage map, we analyzed genetic segregation and quantitative trait loci (QTL) for sesame seed coat color in six generations (P1, P2, F1, BC1, BC2 and F2). Results showed that two major genes with additive-dominant-epistatic effects and polygenes with additive-dominant-epistatic effects were responsible for controlling the seed coat color trait. Average heritability of the major genes in the BC1, BC2 and F2 populations was 89.30%, 24.00%, and 91.11% respectively, while the heritability of polygenes was low in the BC1 (5.43%), in BC2 (0.00%) and in F2 (0.89%) populations. A high-density map was constructed using 724 polymorphic markers. 653 SSR, AFLP and RSAMPL loci were anchored in 14 linkage groups (LG) spanning a total of 1,216.00 cM. The average length of each LG was 86.86 cM and the marker density was 1.86 cM per marker interval. Four QTLs for seed coat color, QTL1-1, QTL11-1, QTL11-2 and QTL13-1, whose heritability ranged from 59.33%–69.89%, were detected in F3 populations using CIM and MCIM methods. Alleles at all QTLs from the black-seeded parent tended to increase the seed coat color. Results from QTLs mapping and classical genetic analysis among the P1, P2, F1, BC1, BC2 and F2 populations were comparatively consistent. This first QTL analysis and high-density genetic linkage map for sesame provided a good foundation for further research on sesame genetics and molecular marker-assisted selection (MAS).

&lt;p&gt;Genetic diversity analysis using molecular markers is an important step for selecting appropriate parents in a soybean breeding program. The aims of this study were to (1) analyze genetic diversity of 29 soybean genotypes assessed with 27 SSR markers for selecting appropriate parents and (2) develop F2 populations to be used for breeding long juvenile (LJ) trait in soybean to&lt;br /&gt;be cultivated in short photoperiod condition. The soybean genotypes used consisted of 11 Indonesian soybean genotypes and 18 genotypes introduced from the USA. F2 populations were developed by crossing Grobogan with three introduced genotypes carrying LJ character. The PIC values of the 27 SSR markers ranged from 0.87 to 0.96. Cluster analysis resulted in three main&lt;br /&gt;clusters at coefficient similarity of 0.76. The five LJ introduced accessions and the nine Indonesian genotypes showed high genetic distances and are useful as parent pairs for developing breeding populations. The F1 progeny phenotypic&lt;br /&gt;performances of the cross far exceeded the performaces of both parents. Three F2 populations were developed by crossing the distantly related soybean genotypes. The F2 populations were verified by using SSR markers and it was found that they segregated in a 1:2:1 ratio confirming the segregation ratio of codominant SSR markers. The F2 populations should be useful for breeding LJ characters to improve soybean productivity in low latitude tropical countries such as Indonesia, which has day length of approximately 12 h all year round.&lt;/p&gt;

DNA markers used in assisting selection method is a safe method in breeding process, due to deletion of environmental conditions, and it is an important tool in preparing linkage map and&nbsp;QTLs mapping.&nbsp;In mulberry silkworm that is, foundation of world sericulture, its major production- economic characteristics are polygenic. In this study, we want to determine QTL(s) affecting cocoon weight trait by AFLP markers. For this reason, we used 20 selected primer combinations from among 81 primers combinations of&nbsp;PstI/TaqI&nbsp;at the level of three F2 populations including 33, 36 and 34 offsprings sample, respectively. These populations were obtained by crossing two lines of Lemon Khorasan (as maternal) and 107 (as paternal). The parental lines, F1 and F2 individuals&rsquo; DNA were extracted with phenol-chloroform method. Then they were digested by two restriction enzymes (TaqI&nbsp;and&nbsp;PstI) and amplified by using of appropriate adaptors. These amplified samples are transfered on annealed 6% polyacrylamide gels. After genotyping of individuals, the linkage maps of populations were drawn by Map manager/QTX and QTL Cartographer ver.2.5 softwares. Number of total and polymorphic bands that formed to 20 primer combinations in each populations were 930, 944, 810 and 142, 171, 178 bands, respectively. Therefore polymorphic frequencies were 15.27, 18.11 and 21.97%. The obtained linkage maps were included in 16, 18 and 24 linkage groups. The total length of this linkage maps and average distance between two markers were 2186.40, 2582.50 and 2392.60 cM, and 18.37, 16.45 and 14.95 cM, respectively. The detection of QTLs numbers of cocoon weight character in each F2 populations also showed 1, 6 and 1 loci in LRS&gt;17 (LOD &gt; 3.7) by compound interval mapping methods, respectively. &nbsp; Key words:&nbsp;Silkworm, QTL analysis, linkage map, molecular markers and AFLP.

Effect of Environment and Genetic Recombination on Subspecies and Economic Trait Differentiation in the F2 and F3 Generations from indica-japonica Hybridization

We obtained a glabrous leaf and hull mutant from a population of radiation mutagenesis of an indica rice cultivar R401. The mutant produced smooth leaves and hairless glumes under normal growth conditions. An F2 population was developed from a cross between a japonica cultivar Nipponbare and the glabrous leaf and hull mutant. By investigating the performance of the F2 population, we found that the mutant phenotype was controlled by a single recessive gene, temporarily designated GLR3. Bulked segregant analysis (BSA) based on the F2 mapping population revealed that GLR3 is located on chromosome 6. By analyzing 417 typical glabrous leaf F2 plants using molecular markers, GLR3 was mapped to a 0.2 cM interval between InDel markers ID27101 and ID27199, and the physical distance between the two markers is 98 kb. Thus we have mapped the gene GLR3, and our work will provide basis for future mechanistic analysis of GLR3 function.