Substoichiometrically Different Mitotypes Coexist in Mitochondrial Genomes of Brassica napus L

Cytoplasmic male sterility (CMS) has been identified in numerous plant species. Brassica napus CMS plants, such as Polima (pol), MI, and Shaan 2A, have been identified independently by different researchers with different materials in conventional breeding processes. How this kind of CMS emerges is unclear. Here, we report the mitochondrial genome sequence of the prevalent mitotype in the most widely used pol-CMS line, which has a length of 223,412 bp and encodes 34 proteins, 3 ribosomal RNAs, and 18 tRNAs, including two near identical copies of trnH. Of these 55 genes, 48 were found to be identical to their equivalents in the “nap” cytoplasm. The nap mitotype carries only one copy of trnH, and the sequences of five of the six remaining genes are highly similar to their equivalents in the pol mitotype. Forty-four open reading frames (ORFs) with unknown function were detected, including two unique to the pol mitotype (orf122 and orf132). At least five rearrangement events are required to account for the structural differences between the pol and nap sequences. The CMS-related orf224 neighboring region (∼5 kb) rearranged twice. PCR profiling based on mitotype-specific primer pairs showed that both mitotypes are present in B. napus cultivars. Quantitative PCR showed that the pol cytoplasm consists mainly of the pol mitotype, and the nap mitotype is the main genome of nap cytoplasm. Large variation in the copy number ratio of mitotypes was found, even among cultivars sharing the same cytoplasm. The coexistence of mitochondrial mitotypes and substoichiometric shifting can explain the emergence of CMS in B. napus

Proteome Dynamics and Physiological Responses to Short-Term Salt Stress in Brassica napus Leaves.

Salt stress limits plant growth and crop productivity and is an increasing threat to agriculture worldwide. In this study, proteomic and physiological responses of Brassica napus leaves under salt stress were investigated. Seedlings under salt treatment showed growth inhibition and photosynthesis reduction. A comparative proteomic analysis of seedling leaves exposed to 200 mM NaCl for 24 h, 48 h and 72 h was conducted. Forty-four protein spots were differentially accumulated upon NaCl treatment and 42 of them were identified, including several novel salt-responsive proteins. To determine the functional roles of these proteins in salt adaptation, their dynamic changes in abundance were analyzed. The results suggested that the up-accumulated proteins, which were associated with protein metabolism, damage repair and defense response, might contribute to the alleviation of the deleterious effect of salt stress on chlorophyll biosynthesis, photosynthesis, energy synthesis and respiration in Brassica napus leaves. This study will lead to a better understanding of the molecular basis of salt stress adaptation in Brassica napus and provides a basis for genetic engineering of plants with improved salt tolerance in the future

Composite Interval Mapping Based on Lattice Design for Error Control May Increase Power of Quantitative Trait Locus Detection.

Experimental error control is very important in quantitative trait locus (QTL) mapping. Although numerous statistical methods have been developed for QTL mapping, a QTL detection model based on an appropriate experimental design that emphasizes error control has not been developed. Lattice design is very suitable for experiments with large sample sizes, which is usually required for accurate mapping of quantitative traits. However, the lack of a QTL mapping method based on lattice design dictates that the arithmetic mean or adjusted mean of each line of observations in the lattice design had to be used as a response variable, resulting in low QTL detection power. As an improvement, we developed a QTL mapping method termed composite interval mapping based on lattice design (CIMLD). In the lattice design, experimental errors are decomposed into random errors and block-within-replication errors. Four levels of block-within-replication errors were simulated to show the power of QTL detection under different error controls. The simulation results showed that the arithmetic mean method, which is equivalent to a method under random complete block design (RCBD), was very sensitive to the size of the block variance and with the increase of block variance, the power of QTL detection decreased from 51.3% to 9.4%. In contrast to the RCBD method, the power of CIMLD and the adjusted mean method did not change for different block variances. The CIMLD method showed 1.2- to 7.6-fold higher power of QTL detection than the arithmetic or adjusted mean methods. Our proposed method was applied to real soybean (Glycine max) data as an example and 10 QTLs for biomass were identified that explained 65.87% of the phenotypic variation, while only three and two QTLs were identified by arithmetic and adjusted mean methods, respectively

Fine Mapping of a Pleiotropic Locus (<i>BnUD1</i>) Responsible for the Up-Curling Leaves and Downward-Pointing Siliques in <i>Brassica napus</i>

Leaves and siliques are important organs associated with dry matter biosynthesis and vegetable oil accumulation in plants. We identified and characterized a novel locus controlling leaf and silique development using the Brassica napus mutant Bnud1, which has downward-pointing siliques and up-curling leaves. The inheritance analysis showed that the up-curling leaf and downward-pointing silique traits are controlled by one dominant locus (BnUD1) in populations derived from NJAU5773 and Zhongshuang 11. The BnUD1 locus was initially mapped to a 3.99 Mb interval on the A05 chromosome with a BC6F2 population by a bulked segregant analysis-sequencing approach. To more precisely map BnUD1, 103 InDel primer pairs uniformly covering the mapping interval and the BC5F3 and BC6F2 populations consisting of 1042 individuals were used to narrow the mapping interval to a 54.84 kb region. The mapping interval included 11 annotated genes. The bioinformatic analysis and gene sequencing data suggested that BnaA05G0157900ZS and BnaA05G0158100ZS may be responsible for the mutant traits. Protein sequence analyses showed that the mutations in the candidate gene BnaA05G0157900ZS altered the encoded PME in the trans-membrane region (G45A), the PMEI domain (G122S), and the pectinesterase domain (G394D). In addition, a 573 bp insertion was detected in the pectinesterase domain of the BnaA05G0157900ZS gene in the Bnud1 mutant. Other primary experiments indicated that the locus responsible for the downward-pointing siliques and up-curling leaves negatively affected the plant height and 1000-seed weight, but it significantly increased the seeds per silique and positively affected photosynthetic efficiency to some extent. Furthermore, plants carrying the BnUD1 locus were compact, implying they may be useful for increasing B. napus planting density. The findings of this study provide an important foundation for future research on the genetic mechanism regulating the dicotyledonous plant growth status, and the Bnud1 plants can be used directly in breeding

Complete mitochondrial genome of Eruca sativa Mill. (Garden rocket).

Eruca sativa (Cruciferae family) is an ancient crop of great economic and agronomic importance. Here, the complete mitochondrial genome of Eruca sativa was sequenced and annotated. The circular molecule is 247,696 bp long, with a G+C content of 45.07%, containing 33 protein-coding genes, three rRNA genes, and 18 tRNA genes. The Eruca sativa mitochondrial genome may be divided into six master circles and four subgenomic molecules via three pairwise large repeats, resulting in a more dynamic structure of the Eruca sativa mtDNA compared with other cruciferous mitotypes. Comparison with the Brassica napus MtDNA revealed that most of the genes with known function are conserved between these two mitotypes except for the ccmFN2 and rrn18 genes, and 27 point mutations were scattered in the 14 protein-coding genes. Evolutionary relationships analysis suggested that Eruca sativa is more closely related to the Brassica species and to Raphanus sativus than to Arabidopsis thaliana

Fine Mapping of a Pleiotropic Locus (BnUD1) Responsible for the Up-Curling Leaves and Downward-Pointing Siliques in Brassica napus

Leaves and siliques are important organs associated with dry matter biosynthesis and vegetable oil accumulation in plants. We identified and characterized a novel locus controlling leaf and silique development using the Brassica napus mutant Bnud1, which has downward-pointing siliques and up-curling leaves. The inheritance analysis showed that the up-curling leaf and downward-pointing silique traits are controlled by one dominant locus (BnUD1) in populations derived from NJAU5773 and Zhongshuang 11. The BnUD1 locus was initially mapped to a 3.99 Mb interval on the A05 chromosome with a BC6F2 population by a bulked segregant analysis-sequencing approach. To more precisely map BnUD1, 103 InDel primer pairs uniformly covering the mapping interval and the BC5F3 and BC6F2 populations consisting of 1042 individuals were used to narrow the mapping interval to a 54.84 kb region. The mapping interval included 11 annotated genes. The bioinformatic analysis and gene sequencing data suggested that BnaA05G0157900ZS and BnaA05G0158100ZS may be responsible for the mutant traits. Protein sequence analyses showed that the mutations in the candidate gene BnaA05G0157900ZS altered the encoded PME in the trans-membrane region (G45A), the PMEI domain (G122S), and the pectinesterase domain (G394D). In addition, a 573 bp insertion was detected in the pectinesterase domain of the BnaA05G0157900ZS gene in the Bnud1 mutant. Other primary experiments indicated that the locus responsible for the downward-pointing siliques and up-curling leaves negatively affected the plant height and 1000-seed weight, but it significantly increased the seeds per silique and positively affected photosynthetic efficiency to some extent. Furthermore, plants carrying the BnUD1 locus were compact, implying they may be useful for increasing B. napus planting density. The findings of this study provide an important foundation for future research on the genetic mechanism regulating the dicotyledonous plant growth status, and the Bnud1 plants can be used directly in breeding

Morphological, photosynthetic and physiological responses of rapeseed leaf to different combinations of red and blue lights at the rosette stage

Rapeseed (Brassica napus L.) is sensitive to light quality. The factory production of rapeseed seedlings for vegetable use and for transplanting in the field requires an investigation of the responses of rapeseed to light quality. This study evaluated the responses of the leaf of rapeseed (cv. ‘Zhongshuang 11’) to different ratios of red-photonflux (RPF) and blue-photonflux (BPF) from light emitting diodes (LEDs). The treatments were set as monochromatic lights, including 100R:0B% and 0R:100B%, and compound lights (CLs), including 75R:25B%, 50R:50B% and 25R:75B%. The total photonflux in all of the treatments was set as 550 μmolm-2s-1. With an increase of BPF, the rapeseed leaves changed from wrinkled blades and down-rolled margins to flat blades and slightly up-rolled margins, and the compact degree of palisade tissue increased. One layer of the cells of palisade tissue was present under 100R:0B%, whereas two layers were present under the other treatments. Compared to 100R:0B%, 0R:100B% enhanced the indexes of leaf thickness, leaf mass per area (LMA), stomatal density, chlorophyll (Chl) content per weight and photosynthetic capacity (Pmax), and the CLs with high BPF ratios enhanced these indexes. However, the 100R:0B% and CLs with high RPF ratios enhanced the net photosynthetic rate (Pn). The leaves under the CLs showed growth vigor, whereas the leaves under 100R:0B% or 0R:100B% were stressed with a low Fv/Fm (photosynthetic maximum quantum yield) and a high content of O2.- and H2O2. The top second leaves under 100R:0B% or 0R:100B% showed stress resistance responses with a high activity of antioxidase, but the top third leaves showed irreversible damage and inactivity of antioxidase. Our results showed that the rapeseed leaves grown under 0R:100B% or CLs with a high BPF ratio showed higher ability to utilize high photonflux, while the leaves grown under 100R:0B% or CLs with a low BPF ratio showed higher efficiency in utilizing low photonflux. Under different R:B photonflux ratios, red and blue lights may play mutual roles in Pn. When the blue light dominated, the Pn showed a B-preference. When the red light dominated, the Pn showed an R-preference. Furthermore, CLs were suitable for the Pn of rapeseed seedlings