Area under disease progress curve and apparent infection rate of Alternaria blight disease of Indian mustard (Brassica juncea) at different plant age

Plant age has a major influence on the incidence of Alternaria blight disease in Indian mustard crops. Disease progression was monitored twice a week on the two chosen Indian mustard cultivars viz., Varuna and Rohini throughout the season. Severity of blight caused by Alternaria brassicae and Alternaria brassicicola decreased with delay in sowing. Calculation for A-value (Area under disease progress curve – AUDPC) and r-value (apparent infection rate) in crops sown on different dates could identify the speed of progress in the disease on leaves and pods, as the crop does not posses resistance to the pathogen till date. Thus, the probable dates of sowing enabling slow disease progress or the weather conditions coinciding with the different crop phenological stages demarcated the advantageous dates of sowing from the disadvantageous ones. However, cultivar Varuna is more susceptible as compared to the other cultivar Rohini, as apparent infection rate both on leaves and pods was higher in former. Highest per cent disease severity (PDS) for season highly correlated with date of sowing, i.e. delayed date of sowing increased PDS.     

β-Aminobutyric Acid-induced Resistance in Brassica juncea against the Necrotrophic Pathogen Alternaria brassicae

β‐Aminobutyric acid (BABA) pretreatment of Brassica plants protected them against the necrotrophic pathogen Alternaria brassicae. The achieved resistance level was much higher than that seen after salicylic acid (SA) and jasmonic acid (JA) pretreatments. BABA pretreatment to the leaves, 1 day before inoculation, led to an inhibition of the oxidative burst and a decrease in SA levels, but did not influence lipoxygenase activity nor cause callose deposition at the site of inoculation. Expression of two marker genes of the SA and JA pathways, namely PR1 and PDF1.2, was enhanced in response to BABA pretreatment. Our results indicate that BABA‐induced resistance is mediated through an enhanced expression of pathogenesis‐related protein genes, independent of SA and JA accumulation.     

Transport and Partitioning of Lead in Indian Mustard (Brassica juncea) and Wheat (Triticum aestivum)

Lead (Pb) contamination in soils is a serious concern because it can be taken up by crops and then transferred through the food chain, posing a potential risk to human health. Indian mustard (Brassica juncea) and wheat (Triticum aestivum) are important crop species known to accumulate heavy metals in their tissues. This study aimed at understanding the transport and accumulation of Pb in these two species and the risk associated with consumption of these foods, which would help us in mitigating accumulation of Pb in edible tissues. The plants were grown at different Pb concentrations for the entire life cycle, and the partitioning of the metal to different tissues was examined. The results showed that plant species differ widely in their ability to transport and accumulate Pb in different tissues. In B. juncea, there was significant accumulation of Pb in both siliques and seeds, whereas most of the Pb in wheat was concentrated in the vegetative tissues and less to the flag leaf and reproductive tissues. In both species, although seed Pb concentrations exceeded acceptable limits, dietary intake did not exceed acceptable limits in most treatments, indicating that more studies on Pb transport and redistribution in crop species is necessary.     

Callus Formation and Morphogenesis of Cultured Mustard (Brassica juncea) Protoplasts

Protoplasts from mustard seedling cotyledons were suspended in Nitsch medium and cultured at 26℃ under low intensity illumination. When colonies were observed in the cultures, fresh medium (3% sucrose replaced mannitol) was added by equalvolume. When protoplasts developed into small callus the cultures were transfered to modified MS agar medium, in which some of them produced roots while an others gave rise to green spots. Shoot-differentiation experiment is in progress.     

提高榨菜离体培养植株再生频率

采用榨菜“浙桐1号”品种为材料,以MS为基本培养基,通过对不同植物生长调节剂的组合和不同外植体等主要因素的筛选,大幅度提高了榨菜离体培养植株再生频率。结果表明,2mg／L6．BA 0．2mg／L2,4-D的组合较为适宜,其不定芽再生频率可达50％,且外植体以下胚轴为好：而CPPU和2,4-D的适宜组合为1．5mg／L 0．2mg／L,其不定芽再生频率高达66．67％,最适外植体为带柄子叶。同时,研究结果显示,添加0．25～1mg／L的GA,对榨菜已分化的不定芽的伸长有抑制作用;子叶柄和下胚轴外植体的分化具有极性现象。     

Chlorophyllase and peroxidase activity during degreening of maturing canola (Brassica napus) and mustard (Brassica juncea) seed

Chlorophyllase and peroxidase activities were measured in relation to seed maturation and degreening in canola ( Brassica napus cvs Westar and Alto) and mustard ( Brassica juncea cvs Cutlass and Lethbridge 22A). Samples of seed collected at the same moisture content were pooled, then divided and used for each assay. During maturation the green pigment (chlorophyll and related pigments) content of canola seed decreased linearly and was lower than that measured in mustard at all moisture contents studied, except for the highest and lowest moisture contents. Chlorophyllides and pheophorbides were detected in canola and were essentially absent in mustard. This difference in accumulation of dephytylated pigments infers differences in the pigment degradation pathways in Brassica species. Interspecific differences in the enzymology of degreening were found. Green pigment degradation was associated with increased chlorophyllase activity and low peroxidase activity in canola and low Chlorophyllase and high perosidase activity in mustard. The possible role of ethylene in seed degreening is discussed.     

Accumulation, detoxification, and genotoxicity of heavy metals in Indian mustard (Brassica juncea L.)

Plants of Indian mustard (Brassica juncea L.) were exposed to different concentrations (15, 30, 60, 120 microM) of (Cd, Cr, Cu, Pb) for 28 and 56 d for accumulation and detoxification studies. Metal accumulation in roots and shoots were analyzed and it was observed that roots accumulated a significant amount of Cd (1980 microg g(-1) dry weight), Cr (1540 microg g(-1) dry weight), Cu (1995 microg g(-1) dry weight), and Pb (2040 microg g(-1) dry weight) after 56 d of exposure, though in shoot this was 1110, 618, 795, and 409 microg g(-1) dry weight of Cd, Cr, Cu, and Pb, respectively. In order to assess detoxification mechanisms, non-protein thiols (NP-SH), glutathione (GSH) and phytochelatins (PCs) were analyzed in plants. An increase in the quantity of NP-SH (9.55), GSH (8.30), and PCs (1.25) micromol g(-1) FW were found at 15 microM of Cd, however, a gradual decline in quantity was observed from 15 microM of Cd onwards, after 56 d of exposure. For genotoxicity in plants, cytogenetic end-points such as mitotic index (MI), micronucleus formation (MN), mitotic aberrations (MA) and chromosome aberrations (CA) were examined in root meristem cells of B. juncea. Exposure of Cd revealed a significant (P < 0.05) inhibition of MI, induction of MA, CA, and MN in the root tips for 24 h. However, cells examined at 24 h post-exposure showed concentration-wise recovery in all the endpoints. The data revealed that Indian mustard could be used as a potential accumulator of Cd, Cr, Cu, and Pb due to a good tolerance mechanisms provided by combined/concerted action of NP-SH, GSH, and PCs. Also, exposure of Cd can cause genotoxic effects in B. juncea L. through chromosomal mutations, MA, and MN formation.     

Development of Plantlets from Leaf Protoplast Culture in Brassica juncea var. tsatsi

Protoplast of two mustard cultivars: Brassica juncea var. tsatsi cv. “Quxian Jiaoercai” and “Bangbangcai”, were isolated by enzymolysis from leaf grown in vitro. Protoplasts were suspended in liquid medium and semi-solidified medium with 0.35% low melting point agarose which formed a thin layer floating on the surface of the liquid medium. The first division appeared after 48h in the culture. One week after the original culture, a diluted medium with gradual dicrease of mannitol concentrations (6%→4%→zero) was then added to the culture three times respectively at one week's interval. In this culture method cell division and formation of microcalli were achieved. During the liquid culture of protoplasts, shaking at 20 rpm from time to time was beneficial in the formation of cell colonies and microcalli. Cell colonies developed into calli of approx 0.5—1mm in diameter one month after culture. The plating efficiency, which defined as the percentage of microcatli to numbers of protoplasts, was 0.2%—1%. Shoot regeneration occured when leaf protoplast-derived calli of “Quxian Jiaoercai” were transferred onto the modified MS medium supplemented with BAP 2.0mg/L, KT 1.0mg/L and NAA 0.2mg/L, and those of -'Bangbangcai" were transferred onto the modified MS medium supplemented with BAP 2.0mg/L. Individual shoot was rooted on a rooting medium supplemented with NAA 0.2 or 0.4 mg/L.     

Phytoremediation of aspirin and tetracycline by Brassica juncea

With the increasing release of pharmaceutical drugs in the environment, research is in progress for investigating alternative methods for their remediation. Various studies have shown the phytoremediation potential of Brassica juncea for metals. The current study was aimed at evaluating the phytoremediation potential of B. juncea for two different pharmaceutical drugs i.e. aspirin and tetracycline in in-vitro conditions. The seeds of B. juncea were germinated and grown for a period of 28 and 24 days for aspirin and tetracycline, respectively. The study analyzed the remediation rate of B. juncea for the selected drugs in three different sets of varying concentration along with any phytotoxic effects exerted by the drugs on the seeds. Preliminary results showed that the average remediation rate of aspirin and tetracycline at the end of experiment was approximately 90% and 71%, respectively. As initial drug concentrations were increased in the media, the remediation rate also improved. However, at higher concentrations, the plants showed phytotoxicity as depicted by the decrease in shoot length of the germinated seeds. These preliminary results indicated that B. juncea could tolerate and remediate pharmaceutical drugs such as analgesics and antibiotics.     

豇豆胰蛋白酶抑制剂基因转化芥菜及抗虫鉴定

用农杆菌介导将豇豆胰蛋白酶抑制剂 (CpTI)基因导入芥菜 ,获得了Kan抗性植株 .经PCR扩增、PCR Southern印迹和Northern印迹分析 ,转化再生植株大部分呈阳性 ,而非转化的再生植株均为阴性 ,证明CpTI基因已存在于芥菜基因组中 .在室内进行了喂虫试验 ,结果表明转基因芥菜抗虫性明显高于对照 ,转基因植株之间存在抗虫性差异     

Assessment of genetic variation within Indian mustard (Brassica juncea) germplasm using random amplified polymorphic DNA markers

Genetic diversity among 45 Indian mustard (Brassica Juncea L.) genotypes comprising 37 germplasm collections, five advance breeding lines and three improved cultivars was investigated at the DNA level using the random amplified polymorphic DNA (RAPD) technique. Fifteen primers used generated a total of 92 RAPD fragments, of which 81 (88%) were polymorphic. Of these, 13 were unique to accession 'Pak85559'. Each primer produced four to nine amplified products with an average of 6.13 bands per primer. Based on pairwise comparisons of RAPD amplification products, Nei and Li's similarity coefficients were calculated to evaluate the relationships among the accessions. Pairwise similarity indices were higher among the oilseed accessions and cultivars showing narrow ranges of 0.77-0.99. An unweighted pair-group method with arithmetic averages cluster analysis based on these genetic similarities placed most of the collections and oilseed cultivars close to each other, showing a low level of polymorphism between the accessions used. However, the clusters formed by oilseed collections and cultivars were comparatively distinct from that of advanced breeding lines. Genetically, all of the accessions were classified into a few major groups and a number of individual accessions. Advanced breeding lines were relatively divergent from the rest of the accessions and formed independent clusters. Clustering of the accessions did not show any pattern of association between the RAPD markers and the collection sites. A low level of genetic variability of oilseed mustard was attributed to the selection for similar traits and horticultural uses. Perhaps close parentage of these accessions further contributed towards their little diversity. The study demonstrated that RAPD is a simple and fast technique to compare the genetic relationship and pattern of variation among the gene pool of this crop.     

Improved microspore culture and doubled-haploid plant regeneration in the brown condiment mustard (Brassica juncea)

 The availability of doubled haploids could greatly contribute to improving seed quality in condiment mustard (Brassica juncea). We have developed an efficient and reliable protocol of microspore culture, modified from that of Baillie et al. (1992), based on a modification of the sucrose concentration of culture media. A comparison of microspore culture media differing in their sucrose content showed that a decrease from 17% (w/v) sucrose during the first 48 h to 10% (w/v) thereafter favoured an increase in the production of embryos whatever the responding genotype tested. Thus, out of the 13 B. juncea genotypes studied, 12 gave rise to embryos, and seven of these embryos could be converted into plants. Doubled-haploid plants were produced after treatment with colchicine. Received: 16 January 2000 / Revision received: 8 August 2000 / Accepted: 20 September 2000     

Involvement of Asada-Halliwell Pathway During Phytoremediation of Chromium (VI) in Brassica juncea L. Plants

Brassica juncea (Indian mustard) L. plants were exposed to different concentrations (0.0, 0.1, 0.3 and 0.5 mM) of Chromium (Cr) and harvested after 30 and 60 days of sowing for the analysis of growth parameters, metal uptake and oxidative stress markers. Significant accumulation of Cr (VI) by B. juncea L. plants resulted in the reduced growth and modulations in the pool of various biochemical stress markers. The toxic effects of Cr (VI) on growth and other stress markers (protein content, lipid peroxidation and antioxidative enzymes viz.SOD, CAT, POD, APOX, GR, DHAR and MDHAR) in B. juncea L. were observed to be concentration and time dependent. Effect of Cr (VI) on biochemical parameters was differential and their maximum activities of SOD, POD, APX, GR, DHAR and lipid peroxidation were recorded at 0.5 mM concentration in 30 days old plants. Whereas, trend in the activities of most of the stress markers was reversed in 60 days old plants. The results obtained from the study suggested that Cr (VI) stress inhibited growth of B. juncea L. plants is directly interrelated with its accumulation and resulted in the modulation in activities of various stress markers.     

Alterations of RNA Editing for the Mitochondrial ATP9 Gene in a New orf220-type Cytoplasmic Male-sterile Line of Stem Mustard （Brassica juncea var. tumida）

RNA editing for the mitochondrlal ATP9 gene of encoding regions has been observed in both cytoplasmic malesterile and maintainer lines of stem mustard, where its editing capacity varied spatially and temporally in the cytoplasmic male sterility （CMS） line. There were four RNA editing sites for the mitochondrial ATP9 gene according to Its normal editing sites in mustard, of which three sites occurred as C-to-U changes and one as a U-to-C change. As a result, the hydrophobicity of deduced ATP9 protein was reduced due to the conversions at its 17th, 45th and 64th positions. Meanwhile, the conservation of deduced ATP9 protein was enhanced by changes at the 56th position. Loss of a specific editing site for ATP9 was observed in juvenile roots, senile roots, senile leaves and floret buds of the CMS line. Comparatively, complete RNA editing for ATP9 gene was retained in juvenile roots, juvenile leaves and floret buds of its maintainer line; however, the loss of a specific editing site for ATP9 gene occurred at senile roots and senile leaves in its maintainer line. These observations allow us to produce a hypothesis that the dysfunction of a specific mitochondrial gene arising from RNA editing could probably be a factor triggering CMS and organ senescence through unknown cross-talk pathways during development.     

影响茎用芥菜愈伤组织诱导和植株再生的因素

茎用芥菜子叶培养在MS 0.25 mg·L-1NAA 0.5 mg·L-16-BA 0.25 mg·L-12,4-D培养基上,可获得较高质量的愈伤组织.愈伤组织培养在MS 1～1 mg·L-16-BA 0.2 mg·L-1NAA培养基上分化频率为8.3%,而在加有羧苄青霉素和头孢霉素的培养基上,最高分化频率可达52.4%.将获得的再生植株转移到1/2MS 0.1 mg·L-1 NAA的生根培养基中,可获得完整植株.     

Evaluation of Microsatellite Markers for Genome Mapping in Indian Mustard (Brassica juncea L)

Microsatellites are robust markers for genome mapping, gene tagging and marker assisted selection. The genus Brassica, having a large and complex genome, requires such type of markers for various applications in genetics and breeding. A set of 202 microsatellite markers were used to screen two parental genotypes of Indian mustard (Brassica juncea) namely, ‘Varuna’, an indigenous cultivar and BEC144, an exotic collection from Poland, of which 36 (17.8%) were informative and usable for segregation analysis. The polymorphic markers detected heterozygosity in advanced generation recombinant inbred lines (RILs) developed earlier from the cross Varuna × BEC144 with a varying frequency that ranged from 0% to 23.5%. Normal Mendelian segregation for majority of microsatellite markers was observed. Eleven markers showed significant deviation from the expected 1:1 segregation ratio. Twelve markers were assigned to six different linkage groups of Indian mustard genome map. The level of polymorphism between the parents and the percentage of useful informative markers as observed in this study, suggested that many more markers are needed to achieve a reasonable coverage of mustard genome. This is the first report on the evaluation of microsatellite markers for genome mapping in B. juncea.     

Alternaria incidence in some alloplasmic lines of Indian mustard (Brassica juncea (L.) Coss.)

Summary The cytoplasmic substitution lines of Brassica juncea (L.) Coss were evaluated for their field resistance to Alternaria blight (Alternaria brassicae). The euplasmic B. juncea cv. RLM 198 had a mesothetic reaction while alloplasmic B. juncea lines with cytoplasms of B. campestris, B. chinensis, and B. japonica were highly susceptible. B. nigra cytoplasm did not have any effect on the disease reaction of the B. juncea genome. However, the alloplasmic lines with the cytoplasm of B. napus and B. carinata revealed a comparatively higher degree of resistance. The study underlined the utility of cytoplasmic manipulations in modifying the phenotypic expression of nuclear genes.     

Ethylene Production during Development of Mustard (Brassica juncea) and Canola (Brassica napus) Seed

An open, continuous flow system was used to investigate ethylene production during degreening of maturing seed of mustard (Brassica juncea cv Cutlass and cv Lethbridge 22A) and canola (Brassica napus cv Westar and cv Alto). Isolated mustard seed evolved higher amounts of ethylene than those of canola, and this was particularly evident both early in embryogeny and later during the desiccation phase of seed maturation. The silique walls produced negligible amounts of ethylene in both species. The concentrations of ethylene surrounding seed as they matured within siliques were significantly higher in mustard than in canola, and this interspecies difference was greatest during the seed desiccation phase. In mustard, a 4-fold increase in silique internal ethylene levels was apparent during desiccation. In comparison, only a moderate increase in silique-derived ethylene occurred in canola.