Rapid Cycling Brassica Rapa Research Articles

Adventitious shoot regeneration from cotyledonary explants of rapid-cycling fast plants of Brassica rapa L

Rapid-cycling fast plants (Brassica rapa; RCBr) is also known as Wisconsin Fast Plant and is widely used in K-12 and undergraduate studies. RCBr has a short generation time (seed-to-seed in 30–60 days), which allows for the completion of experiments in a semester. Previous studies have shown that cotyledonary explants with attached petioles are capable of generating shoots. However, there is no published adventitious shoot regeneration protocol to date. Sterile cotyledonary explants were excised; all edges and petioles were removed. Five-day-old cotyledonary explants produced shoots on a Murashige and Skoog medium containing 1.5 mg/L thiadiazuron (TDZ) and 0.5 mg/L 1-naphthaleneacetic acid (NAA) (FPM I) at a mean rate of 8.8%. This rate increased to 14.8% in explants placed on FPM I medium supplemented with 5.0 mg/L silver nitrate (AgNO3) (SRM 2). The rate increased to 32.5% when 5-day-old explants, excised from the part of the cotyledon nearest to the petiole, were placed adaxial side up on SRM 2 medium. The shoot regeneration rate increased to 44.5% using 4-day-old cotyledonary explants. A shoot regeneration rate of 23% was observed among 9-day-old leaf explants. Shoots from cotyledonary explants were elongated on basal medium with 0.5 mg/L NAA, rooted on basal medium, and later acclimatized. This is the first report of shoot regeneration from cotyledonary explants of rapid-cycling Brassica rapa without pre-existing meristematic tissues.

Identification of sources and inheritance of resistance of Chinese Brassica vegetables to white blister

AbstractThere is no information in the literature about the variability of resistance of Chinese Brassica vegetables, pak choi and Chinese cabbage, to the white blister disease caused by Albugo candida (Pers.) Kuntze. A collection of 43 accessions of pak choi and 19 accessions of Chinese cabbage was screened for resistance to the Portuguese A. candida isolate Ac 506 at the cotyledon stage. Different levels of resistance were found among the germplasm tested, ranging from complete resistance to full susceptibility. Most of the accessions were highly susceptible with less than 10% of resistant seedlings, and only four accessions of pak choi, presenting more than 50% resistant seedlings, were considered as new sources or resistance to white blister. Inheritance of resistance at the cotyledon stage was studied in two crosses between the most resistant pak choi accession, BRA 117, and the highly susceptible rapid cycling Brassica rapa line CrGC 1.19. It was proposed that resistance to white blister in pak choi BRA 117 is controlled by two nuclear genes with dominant recessive epistatic gene action.

Development of public immortal mapping populations, molecular markers and linkage maps for rapid cycling Brassica rapa and B. oleracea

Publicly available genomic tools help researchers integrate information and make new discoveries. In this paper, we describe the development of immortal mapping populations of rapid cycling, self-compatible lines, molecular markers, and linkage maps for Brassica rapa and B. oleracea and make the data and germplasm available to the Brassica research community. The B. rapa population consists of 160 recombinant inbred (RI) lines derived from the cross of highly inbred lines of rapid cycling and yellow sarson B. rapa. The B. oleracea population consists of 155 double haploid (DH) lines derived from an F1 cross between two DH lines, rapid cycling and broccoli. A total of 120 RFLP probes, 146 SSR markers, and one phenotypic trait (flower color) were used to construct genetic linkage maps for both species. The B. rapa map consists of 224 molecular markers distributed along 10 linkage groups (A1-A10) with a total distance of 1125.3 cM and a marker density of 5.7 cM/marker. The B. oleracea genetic map consists of 279 molecular markers and one phenotypic marker distributed along nine linkage groups (C1-C9) with a total distance of 891.4 cM and a marker density of 3.2 cM/marker. A syntenic analysis with Arabidopsis thaliana identified collinear genomic blocks that are in agreement with previous studies, reinforcing the idea of conserved chromosomal regions across the Brassicaceae.

Development and Application of Inquiry Module Regarding Plant Responses to Light and Gravity Using Rapid-cycling Brassica rapa

Development and Application of Inquiry Module Regarding Plant Responses to Light and Gravity Using Rapid-cycling Brassica rapa

Introducing a Warm Temperature Adapted Rapid Cycling “Brassica Rapa” to a Thai Science Classroom: A Preliminary Study on Students’ Involvement in Realistic Investigation

AbstractA warm-temperature-adapted breeding pool of Wisconsin Fast Plants (rapid-cycling Brassica rapa; RBr) was developed as a biologically instructional material. Experimentation with the warm-temperature-adapted RBr was used to study students’ investigative skills, understanding of the nature of science, and knowledge of the life cycle of flowering plants. Three secondary school students were involved in this realistic investigation. Students were provided with information on the warm-temperature-adapted RBr from previous experiments. The students’ knowledge of flowering plants and the Views of Scientific Inquiry Questionnaire (VOSI) were used to investigate participants’ understanding of the nature of science. Students had an opportunity to participate in every step of the investigation process including asking and refining questions, finding information, planning and designing, collecting and analyzing data, drawing conclusions, and communicating results. The results demonstrated that RBr adapt well to warm temperatures. The plants were able to grow at warm temperatures ranging from 28-32˚C. The students’ investigative skills improved in terms of asking questions, making predictions, collecting data, and communicating results. Their knowledge of plant life also improved. However, the students’ understanding of the nature of science was not affected.

Application Effects of Inquiry Modules to Study Environmental Pollution Using Rapid-cycling Brassica rapa for the Elementary School Students

Application Effects of Inquiry Modules to Study Environmental Pollution Using Rapid-cycling Brassica rapa for the Elementary School Students

Mapping the Anthocyaninless (anl) Locus in Rapid-Cycling Brassica rapa (RBr) to Linkage Group R9

BackgroundAnthocyanins are flavonoid pigments that are responsible for purple coloration in the stems and leaves of a variety of plant species. Anthocyaninless (anl) mutants of Brassica rapa fail to produce anthocyanin pigments. In rapid-cycling Brassica rapa, also known as Wisconsin Fast Plants, the anthocyaninless trait, also called non-purple stem, is widely used as a model recessive trait for teaching genetics. Although anthocyanin genes have been mapped in other plants such as Arabidopsis thaliana, the anl locus has not been mapped in any Brassica species.ResultsWe tested primer pairs known to amplify microsatellites in Brassicas and identified 37 that amplified a product in rapid-cycling Brassica rapa. We then developed three-generation pedigrees to assess linkage between the microsatellite markers and anl. 22 of the markers that we tested were polymorphic in our crosses. Based on 177 F2 offspring, we identified three markers linked to anl with LOD scores ≥ 5.0, forming a linkage group spanning 46.9 cM. Because one of these markers has been assigned to a known B. rapa linkage group, we can now assign the anl locus to B. rapa linkage group R9.ConclusionThis study is the first to identify the chromosomal location of an anthocyanin pigment gene among the Brassicas. It also connects a classical mutant frequently used in genetics education with molecular markers and a known chromosomal location.

Effects of nutrient and CO2 availability on tolerance to herbivory in Brassica rapa

The ability of plants to recover from herbivore damage and maintain their fitness depends on physiological mechanisms that are affected by the availability of resources such as carbon and soil nutrients. In this study, we explored the effects of increased carbon and nutrient availability on the response of rapid cycling Brassica rapa to damage by the generalist herbivore, Trichoplusia ni (Noctuidae), in a greenhouse experiment. Using fruit mass as an estimate of plant fitness, we tested three physiological models, which predict either an increase or a decrease of tolerance to herbivory with increasing resource availability. We used leaf demography to examine some plausible mechanisms through which resource availability may affect tolerance. Our results contra- dict all models, and, rather, they support a more complicated view of the plasticity of resource uptake and allocation than the ones considered by the models tested. Fruit mass was negatively affected by herbi- vore damage only under elevated CO2, and only for certain harvest dates. Increased CO2 had no effect on the number of leaf births, but it decreased leaf longevity and the total number of leaves on a plant. Nutrient addition increased the number of leaf births, leaf longevity and the total number of leaves on a plant. We conclude that a shortening of the life span of the plants, brought about by elevated CO2, was responsible for a higher susceptibility of plants to herbivore damage under high CO2 concentration.

Teaching Human Genetics with Mustard: Rapid Cycling Brassica rapa (Fast Plants Type) as a Model for Human Genetics in the Classroom Laboratory

We have developed experiments and materials to model human genetics using rapid cycling Brassica rapa, also known as Fast Plants. Because of their self-incompatibility for pollination and the genetic diversity within strains, B. rapa can serve as a relevant model for human genetics in teaching laboratory experiments. The experiment presented here is a paternity exclusion project in which a child is born with a known mother but two possible alleged fathers. Students use DNA markers (microsatellites) to perform paternity exclusion on these subjects. Realistic DNA marker analysis can be challenging to implement within the limitations of an instructional lab, but we have optimized the experimental methods to work in a teaching lab environment and to maximize the "hands-on" experience for the students. The genetic individuality of each B. rapa plant, revealed by analysis of polymorphic microsatellite markers, means that each time students perform this project, they obtain unique results that foster independent thinking in the process of data interpretation.

Does Herbicide Resistance Have a Cost in Brassica rapa?

The evolution of resistance to pesticides has become a classic example of natural selection at work (Palumbi, 2001). As humans chemically modify their environment, organisms with short generation times respond. Examples include resistance of bacteria to antibiotics, of insects to insecticides, and of weeds to herbicides. Here the focus is on resistance to herbicides in weedy plants. In particular, genetic strains of rapid-cycling Brassica rapa offer opportunities to study resistance to the herbicide atrazine, a triazine herbicide. In the presence of the herbicide, individuals of the resistant strain survive whereas individuals of the susceptible strain die. This experiment has been used in a college-level biology class but may, also be applicable to an Advanced Placement Biology class in high school. In addition to the biological content, students gain experience in hypothesis formation and testing, experimental design, and statistical analysis. Herbicide resistance may come with a cost to the organism. Resistance to triazine herbicides is often dependent on a mutation that also inhibits photosynthesis (Warwick, 1991; Holt et al., 1993). For atrazine to kill a plant, it first binds to the polypeptide psbA, located in the thylacoid membrane of chloroplasts. In resistant plants of Amaranthus hybridus, the herbicide fails to bind to psbA. One mutation, in the chloroplast DNA coding for psbA, accounts for the difference between susceptible and resistant plants (Hirschberg & McIntosh, 1983). The mutation that confers resistance also inhibits electron transfer in Photosystem II and under certain environmental conditions will inhibit the rate of photosynthesis (Dekker & Sharkey, 1992). Photosystem II is part of the energy-capturing apparatus of chloroplasts. When chlorophyll in Photosystem II absorbs a photon of light, it initiates a complex cascade of electron transport that eventually captures the energy in chemical form in the chloroplast. If electron transport is inhibited, less energy, can be captured (see Campbell & Reece, 2002, for a lucid and detailed explanation). In the absence of the herbicide, resistant organisms may be disadvantaged. Reduction of photosynthesis will lead to reduced biomass accumulation and may negatively affect competitive ability and fitness (Warwick, 1991; Warwick & Black, 1994). Williams et al. (1995) worked with two strains of jimsonweed (Datura stramonium) that were either susceptible or resistant to the triazine herbicides. They found that resistant jimsonweeds, when grown alone, had lower total biomass and lower reproductive biomass than susceptible jimsonweeds. When jimsonweed was grown in competition with maize, its total biomass and reproductive biomass were still lower than without competition. Resistant jimsonweeds were more negatively affected by competition than susceptible jimsonweeds. Cost of resistance is defined as the reduction in fitness of resistant plants grown in the absence of herbicide. Brassica rapa, in the form of Wisconsin Fast Plants[TM] (Hafner, 1990), is ideal for studying the cost of herbicide resistance. An atrazine-resistant strain (Aaa, Rci, zr, atrazine resistant) can be grown alone and in combination with an atrazine-susceptible strain without the use of herbicide. Cost of resistance can be measured as the reduction in growth and/or reproduction of the resistant strain relative to the susceptible strain. Competitive ability can be measured by comparing plants of one strain grown alone with plants of the same strain grown in combination with the second strain. When two plants of the same strain are grown together they experience intrastrain competition, whereas a resistant plant and a susceptible plant growing together experience inter-strain competition. Based on the experiment described below, students should be able to answer the questions: * Does herbicide resistance have a cost in terms of growth and reproduction in rapid-cycling Brassica rapa? …

Genetic Variation and Selection Response in Model Breeding Populations of Brassica rapa Following a Diversity Bottleneck

Domestication and breeding share a common feature of population bottlenecks followed by significant genetic gain. To date, no crop models for investigating the evolution of genetic variance, selection response, and population diversity following bottlenecks have been developed. We developed a model artificial selection system in the laboratory using rapid-cycling Brassica rapa. Responses to 10 cycles of recurrent selection for cotyledon size were compared across a broad population founded with 200 individuals, three bottleneck populations initiated with two individuals each, and unselected controls. Additive genetic variance and heritability were significantly larger in the bottleneck populations prior to selection and this corresponded to a heightened response of bottleneck populations during the first three cycles. However, the overall response was ultimately greater and more sustained in the broad population. AFLP marker analyses revealed the pattern and extent of population subdivision were unaffected by a bottleneck even though the diversity retained in a selection population was significantly limited. Rapid gain in genetically more uniform bottlenecked populations, particularly in the short term, may offer an explanation for why domesticators and breeders have realized significant selection progress over relatively short time periods.

Characterizing genotype specific differences in survival, growth, and reproduction for field grown, rapid cycling Brassica rapa

Rapid cycling Brassica rapa (RCBr) develops rapidly, and has both small adult size and a brief life cycle. Yet, in spite of many investigations using RCBr, extremely few plant ecologists have used this plant in the field. This study is the first to describe the genotype specific variation in traits describing survival, growth, and reproduction for field grown, RCBr. I also identify traits associated with fitness. Five genotypes of RCBr were used: standard, anthocyaninless, yellow-green, anthocyaninless and hairless, and anthocyaninless and yellow-green. Plants were grown outside in a “common garden”. Eight plant traits were measured: life span, height, growth rate, leaf size, number of flowers and fruits, fruit set, and fitness. All traits, except life span, differed significantly among the five plant genotypes. Correlation analysis revealed that fitness increased as each of these of seven plant traits increased. This study demonstrates that RCBr can serve as a model organism in ecological field studies.

Zinc Availability in Hydroponic Culture Influences Glucosinolate Concentrations in Brassica rapa

Rapid cycling Brassica rapa L. were grown for 7 days in the presence of 11 levels of zinc (Zn) in hydroponic solution culture and evaluated for changes in Zn and glucosinolate (GS) content. Zinc levels were 0.05, 1, 5, 10, 25, 50, 75, 100, 125, 150, and 200 mg·L-1 Zn. Plants grown in solutions with ≥50 mg·L-1 Zn displayed severe Zn toxicity symptoms, grew little, or died and were not subsequently evaluated for GS content. Shoot Zn concentrations increased linearly with increasing Zn treatment levels. Gluconapin, which accounted for nearly 90% of the aliphatic GSs present, was the only aliphatic GS influenced by Zn, and decreased linearly with increasing Zn levels. Accumulation of glucobrassicin and 4-methoxyglucosbrassicin, both indole GSs, responded with a linear increase and quadratically, respectively, to Zn fertility. An aromatic GS, gluconasturtiin, was also influenced by Zn levels in solution, and had a quadratic response to increasing Zn. This suggested that Zn fertility can influence changes in GS that may affect flavor (bitterness, etc.) or medicinal attributes associated with the GS and their breakdown products, as well as elevate the nutritional status of Zn in the leaves of Brassica.

A Method to Teach Age-Specific Demography with Field Grown Rapid Cycling Brassica rapa (Wisconsin Fast Plants)

A Method to Teach Age-Specific Demography with Field Grown Rapid Cycling <i>Brassica rapa</i> (Wisconsin Fast Plants)

048 Effects of Inbreeding in a “Wisconsin Fast Plants” Brassica rapa Population

Wisconsin Fast Plants (WFP) are small, rapid-cycling Brassica rapa populations that were developed by Paul Williams in the Dept. of Plant Pathology at the Univ. of Wisconsin, to facilitate classroom demonstration of biological principals. WFP exist as heterogenous populations, which have been selected for expression of different mutant phenotypes. Because of self-incompatibility mechanisms, it has been difficult to develop inbred lines of WFP via self-feritilization. Our objective was to inbreed a WFP population through full-sib mating. Genetic diversity was calculated for eight individuals from each of eight different WFF population, using 69 polymorphic RAPD (molecular marker) bands. The eight different WPF populatons were randomly mated (via chain crossing) for two generations. Six cycles of full-sib mating were initiated on 130 random families. After six cycles of full-sib mating, 79 families remain. The loss of families, during the process of inbreeding, may have been due to selection or drift. However, the expectation is that genetic variance will increase. The 79 inbred families express an array of different WFP phenotypes, e.g., anthocyanin pigmentation, yellow cotyledon, plant height, and seed color at different combinations in different inbred lines.

Exploring Cost Constraints on Stem Elongation in Plants Using Phenotypic Manipulation

Exploring Cost Constraints on Stem Elongation in Plants Using Phenotypic Manipulation

Realized Defense of Artificially Selected Lines of Brassica rapa: Effects of Quantitative Genetic Variation in Foliar Glucosinolate Concentration

Herbivore species-specific responses to a plant secondary compound could result in variable selection on the concentration of that plant chemical. The purpose of this study was to determine if lines of a rapid-cycling Brassica rapa (L.) that had been artificially selected for quantitative differences in foliar glucosinolate content also differed in their realized defense , defined as the number of eggs laid on or level of damage incurred by a plant when exposed to herbivores. I asked whether defense was conferred at the oviposition or feeding stage, whether 2 herbivore species respond similarly to different foliar glucosinolate concentrations, and whether these same 2 species could impose selection in different directions on foliar glucosinolate concentration. I used an oligophagous herbivore, Pieris rapae (L.), and a polyphagous herbivore, Trichoplusia ni Hübner. Feeding was examined for both herbivore species, and oviposition was examined for P. rapae . P. rapae adult females did not favor any single selection line over another as a site for oviposition. However, P. rapae larvae did remove greater amounts of leaf area on plants from lines selected for low glucosinolate content when given a choice. In addition, T. ni larvae caused significantly greater amounts of damage on low glucosinolate line plants. Thus, quantitative variation in glucosinolate content among selected lines does confer differences in realized defense against these 2 herbivores. However, because P. rapae adult females do not appear to discriminate among plants that differ in glucosinolate content, defense is conferred at the level of feeding. Further, in contrast to previous studies, larvae of both the polyphagous and the oligophagous herbivore species responded similarly to quantitative variation in secondary compounds, causing more damage on low glucosinolate lines. This parallel response suggests that both herbivore species should impose selection on defense in the same direction in B. rapa for increased levels of glucosinolate production.

One step beyond lethal equivalents: characterization of deleterious loci in the rapid cycling Brassica rapa L. base population.

The total number of lethal equivalents as defined by Morton, Crow and Muller (1956) is a function of three parameters: M, the number of loci at which deleterious mutations can occur, q, the frequency of the deleterious alleles at each locus, and s, their selective value. A new approach based on multi-generation inbreeding data is outlined and used to infer these three parameters as well as the dominance coefficient, h, in a self-incompatible species, Brassica rapa L. Germination and flowering data from thirty bud-selfed lines of fast-cycling B.rapa were assessed over three generations. Germination and flowering were significantly postponed by inbreeding but germination and flowering success were not so strongly decreased. Estimates of the average s values were obtained but it was not possible to get separate estimates of M and q. For both characters, the average dominance coefficient was particularly low. The number of lethal equivalents at the zygotic level was around two for germination and three for flowering, which, owing to the self-incompatibility of B.rapa, is an unexpectedly low value. These results may be explained by past biparental inbreeding which in turn may have increased self-compatibility thus allowing the purging of more deleterious alleles than under strict self-incompatibility.

Direct shoot formation and plant regeneration from cotyledon explants of rapid-cycling Brassica rapa

An in vitro culture system for direct shoot regeneration from cotyledon explants of rapid-cycling Brassica rapa was developed. Cotyledons from 3-d-old seedlings, when cultured on Murashige and Skoog (MS) medium supplemented with 20 µM N6-benzyladenine (BA) and 2 µM α-naphthaleneacetic acid (NAA), regenerated shoots directly at a frequency of 20%. The addition of 2 µM aminoethoxyvinylglycine (AVG) to this medium increased shoot regeneration to 33%, but silver nitrate drastically inhibited shoot regeneration. Shoot regeneration occurred directly, at the petiolar cut ends of cotyledonary explants, between 10 to 17 d in culture. The highest percentage of regeneration (33%) was obtained from 3-d-old seedlings. NAA was the most effective auxin for root induction and development, with 49% of shoots producing roots after 2 wk on medium containing 1.0 µM NAA. Regenerated plantlets were grown to maturity in pots containing peat moss and vermiculite (1:1). These plants were morphologically normal and fertile. With this protocol, over 100 independently derived, flowering R0 plants were obtained from 40 regenerating cotyledonary explants within 40 d after culture initiation.

Convergent pathways of gibberellin A1 biosynthesis in Brassica

[2H, 3H]Gibberellin A4 (GA4) or [2H, 3H] GA9 were applied to the shoot tips of seedlings of elongated internode (ein), a tall mutant of rapid cycling Brassica rapa. Following [2H]GA9 application, [2H]GA51, [2H]GA20 and [2H]GA4 were identified as products by GC-MS, while [2H]GA34 and [2H]GA1 were formed from [2H]GA4. Other isotopically labelled products, including abundant putative conjugates, were also produced, but were not identified. Thus, in B. rapa, GA1 biosynthesis involves the convergence of at least two metabolic pathways; it can be formed via GA4 or GA20, the latter of which can originate from GA9 or from GA19.