BnaC7.ROT3, the causal gene of cqSL-C7, mediates silique length by affecting cell elongation in Brassica napus.

Siliques are a major carbohydrate source of energy for later seed development in rapeseed (Brassica napus). Thus, silique length has received great attention from breeders. We previously detected a novel quantitative trait locus cqSL-C7 that controls silique length in B. napus. Here, we further validated the cqSL-C7 locus and isolated its causal gene (BnaC7.ROT3) by map-based cloning. In 'Zhongshuang11' (parent line with long siliques), BnaC7.ROT3 encodes the potential cytochrome P450 monooxygenase CYP90C1, whereas in 'G120' (parent line with short siliques), a single nucleotide deletion in the fifth exon of BnaC7.ROT3 results in a loss-of-function truncated protein. Sub-cellular localization and expression pattern analysis revealed that BnaC7.ROT3 is a membrane-localized protein mainly expressed in leaves, flowers and siliques. Cytological observations showed that the cells in silique walls of BnaC7.ROT3-transformed positive plants were longer than those of transgene-negative plants in the background of 'G120', suggesting that BnaC7.ROT3 affects cell elongation. Haplotype analysis demonstrated that most alleles of BnaC7.ROT3 are favorable in B. napus germplasms, and its homologs may also be involved in silique length regulation. Our findings provide novel insights into the regulatory mechanisms of natural silique length variations and valuable genetic resources for the improvement of silique length in rapeseed.

Optimizing glyphosate tolerance in rapeseed by CRISPR/Cas9-based geminiviral donor DNA replicon system with Csy4-based single-guide RNA processing.

Rapeseed (Brassica napus L.) is an important oil crop worldwide, and effective weed control can protect its yield and quality. Farmers can benefit from cultivars tolerant to herbicides such as glyphosate. Amino acid substitutions in enolpyruvylshikimate-3-phosphate synthase (EPSPS) render the plant less sensitive to glyphosate. Therefore, we aimed to optimize the glyphosate tolerance trait in rapeseed via endogenous EPSPS modification. To achieve effective gene replacement in B. napus L., we employed a CRISPR/Cas9 system expressing single-guide RNAs (sgRNAs) cleaved by the CRISPR-associated RNA endoribonuclease Csy4 from Pseudomonas aeruginosa, for targeted induction of double-strand breaks. Both the donor template and a geminiviral replicon harbouring an sgRNA expression cassette were introduced into plant cells. Using sgRNAs targeting adjacent donor DNA template containing synonymous mutations in sgRNA sites, we achieved precise gene replacements in the endogenous B. napus EPSPS gene, BnaC04EPSPS, resulting in amino acid substitutions at frequencies up to 20%. Rapeseed seedlings harbouring these substitutions were glyphosate-tolerant. Furthermore, modifications in BnaC04EPSPS were precisely transmitted to the next generation. Our genome editing strategy enables highly efficient gene targeting and the induction of glyphosate tolerance in oilseed rape.

Interaction between Brassica napus polygalacturonase inhibition proteins and Sclerotinia sclerotiorum polygalacturonase: implications for rapeseed resistance to fungal infection.

MAIN CONCLUSION: BnPGIPs interacted with Sclerotinia sclerotiorum PGs to improve rapeseed SSR resistance at different levels; the BnPGIP-overexpression lines did not affect plant morphology or seed quality traits. Plant polygalacturonase-inhibiting proteins (PGIPs) play a crucial role in plant defence against phytopathogenic fungi by inhibiting fungal polygalacturonase (PG) activity. We overexpressed BnPGIP2, BnPGIP5, and BnPGIP10 genes in an inbred line 7492 of rapeseed (Brassica napus). Compared with 7492WT, the overexpression of BnPGIP2 lines significantly increased Sclerotinia sclerotiorum resistance in both seedlings and adult plants. BnPGIP5 overexpression lines exhibited decreased S. sclerotiorum disease symptoms in seedlings only, whereas BnPGIP10 overexpression lines did not improve Sclerotinia resistance for seedlings or adult plants. Quantitative real-time PCR analysis of S. sclerotiorum PG1, SsPG3, SsPG5, and SsPG6 genes in overexpressing BnPGIP lines showed that these pathogenic genes in the Sclerotinia resistance transgenic lines exhibited low expression in stem tissues. Split-luciferase complementation experiments confirmed the following: BnPGIP2 interacts with SsPG1 and SsPG6 but not with SsPG3 or SsPG5; BnPGIP5 interacts with SsPG3 and SsPG6 but not with SsPG1 or SsPG5; and BnPGIP10 interacts with SsPG1 but not SsPG3, SsPG5, or SsPG6. Leaf crude protein extracts from BnPGIP2 and BnPGIP5 transgenic lines displayed high inhibitory activity against the SsPG crude protein. BnPGIP-overexpression lines with Sclerotinia resistance displayed a lower accumulation of H2O2 and higher expression of the H2O2-removing gene BnAPX (ascorbate peroxidase) than 7492WT, as well as elevated expression of defence response genes including jasmonic acid/ethylene and salicylic acid pathways after S. sclerotiorum infection. The plants overexpressing BnPGIP exhibited no difference in either agronomic traits or grain yield from 7492WT. This study provides potential target genes for developing S. sclerotiorum resistance in rapeseed.

Heterosis Derived From Nonadditive Effects of the BnFLC Homologs Coordinates Early Flowering and High Yield in Rapeseed (Brassica napus L.).

Early flowering facilitates crops to adapt multiple cropping systems or growing regions with a short frost-free season; however, it usually brings an obvious yield loss. In this study, we identified that the three genes, namely, BnFLC.A2, BnFLC.C2, and BnFLC.A3b, are the major determinants for the flowering time (FT) variation of two elite rapeseed (Brassica napus L.) accessions, i.e., 616A and R11. The early-flowering alleles (i.e., Bnflc.a2 and Bnflc.c2) and late-flowering allele (i.e., BnFLC.A3b) from R11 were introgressed into the recipient parent 616A through a breeding strategy of marker-assisted backcross, giving rise to eight homozygous near-isogenic lines (NILs) associated with these three loci and 19 NIL hybrids produced by the mutual crossing of these NILs. Phenotypic investigations showed that NILs displayed significant variations in both FT and plant yield (PY). Notably, genetic analysis indicated that BnFLC.A2, BnFLC.C2, and BnFLC.A3b have additive effects of 1.446, 1.365, and 1.361 g on PY, respectively, while their dominant effects reached 3.504, 2.991, and 3.284 g, respectively, indicating that the yield loss caused by early flowering can be successfully compensated by exploring the heterosis of FT genes in the hybrid NILs. Moreover, we further validated that the heterosis of FT genes in PY was also effective in non-NIL hybrids. The results demonstrate that the exploration of the potential heterosis underlying the FT genes can coordinate early flowering (maturation) and high yield in rapeseed (B. napus L.), providing an effective strategy for early flowering breeding in crops.

Clinical Efficacy of Laparoscopic Surgery for T4 Colon Cancer Compared with Open Surgery: A Single Center's Experience.

BACKGROUND: Laparoscopic surgery for T4 colon cancer remains controversial according to many colorectal cancer guidelines. The aim of this study was to compare short- and long-term outcomes in patients who underwent T4 colon cancer resection by laparoscopy versus open surgery. METHODS: Patients who underwent T4 colon cancer resection either by laparoscopy or by open surgery between January 2012 and January 2017 were included and used to perform a retrospective cohort analysis. Demographics, patient characteristics, short-term outcomes, and long-term oncological outcomes were compared between two groups. Multivariate analyses were used to define prognostic factors of overall survival. RESULTS: Groups were comparable in terms of preoperative characteristics and demographics. Intraoperative blood loss (127.3 versus 226.1 mL, P = .001) and hospital stay (11.6 versus 14.8 days, P = .001) were significantly reduced in the laparoscopic group compared with the open group. Operative time, bowel movement, time to soft diet, and lymph nodes harvested did not significantly differ between the two groups. R0 resection achieved 100% in both the groups. Similarly, the overall survival rate and disease-free survival rate in stage II and stage III disease showed no significant differences. Multivariate analyses showed that intraoperative blood loss was a significantly independent factor related to a poor prognosis. CONCLUSIONS: This study suggests that laparoscopy for T4 colon cancer can be safely performed with superior short-term outcomes, such as less intraoperative blood loss and shorter time of hospital stay compared with open surgery, and with similar long-term oncological outcomes. Therefore, laparoscopic procedure could be a viable option in selected patients.

Single-incision laparoscopic versus conventional laparoscopic right colectomy: A systematic review and meta-analysis.

BACKGROUND: Single-incision laparoscopic surgery has gained widespread attention because of its potential benefits such as less skin incision and faster recovery. Up to now, only one meta-analysis (performed in 2013; including 9 studies, a total of 585 cases) compared single-incision laparoscopic right colectomy (SILRC) with conventional laparoscopic right colectomy (CLRC). An updated meta-analysis was undertaken to explore more convinced comparative findings between SILRC and CLRC. MATERIALS AND METHODS: The search for studies that compared SILRC with CLRC was done on PubMed, Embase, Web of Science, and the Cochrane Library. A total of 17 studies (including 1778 cases) were identified, the data of appointed outcomes were extracted and analyzed. RESULTS: Patient demographics (age, gender, body mass index and previous abdominal operation) did not differ significantly. No significant differences were found between SILRC and CLRC in operative time, conversion, reoperation, perioperative complications, postoperative mortality, and 30-days readmission. Pathological outcomes, including lymph nodes harvested, proximal resection margin, and distal resection margin, were similar. SILRC showed less estimated blood loss (weighted mean difference [WMD]: -15.67 ml; 95% confidence interval [CI], -24.36 to -6.98; p = 0.0004), less skin incisions (WMD: -1.56 cm; 95%CI, -2.63 to -0.49; p = 0.004) and shorter hospital stay (WMD: -0.73d; 95%CI, -1.04 to -0.41; p < 0.00001), without publication bias. CONCLUSION: SILRC may provide a safe and feasible alternative to CLRC with similar short-term outcomes and aesthetic advantage of less skin incision. Well-designed randomized controlled trials, involving large cases and carrying long-term outcomes, are needed.

Host-parasite coevolution in populations of constant and variable size.

BACKGROUND: The matching-allele and gene-for-gene models are widely used in mathematical approaches that study the dynamics of host-parasite interactions. Agrawal and Lively (Evolutionary Ecology Research 4:79-90, 2002) captured these two models in a single framework and numerically explored the associated time discrete dynamics of allele frequencies. RESULTS: Here, we present a detailed analytical investigation of this unifying framework in continuous time and provide a generalization. We extend the model to take into account changing population sizes, which result from the antagonistic nature of the interaction and follow the Lotka-Volterra equations. Under this extension, the population dynamics become most complex as the model moves away from pure matching-allele and becomes more gene-for-gene-like. While the population densities oscillate with a single oscillation frequency in the pure matching-allele model, a second oscillation frequency arises under gene-for-gene-like conditions. These observations hold for general interaction parameters and allow to infer generic patterns of the dynamics. CONCLUSION: Our results suggest that experimentally inferred dynamical patterns of host-parasite coevolution should typically be much more complex than the popular illustrations of Red Queen dynamics. A single parasite that infects more than one host can substantially alter the cyclic dynamics.

Life histories and Cope's rule from an explicit resource-consumer model based on metabolic theory.

We explore the consequences of metabolic theory for life histories and life history evolution. We use a mathematical model for an iteroparous species and its resources, taking into account the allometric scaling of consumption, metabolism, and mortality with consumer body mass. Mortality is assumed to be density-dependent, and the dynamics of resources are modeled explicitly. By evaluating life history features in equilibrium populations, we find that in populations that use more or faster growing resources the individuals have a shorter lifespan and a higher mortality, and that individuals in populations with a larger adult body mass have a longer lifespan, a larger number of offspring per female, and a higher biomass density. When we allow the adult body mass to evolve, it increases in time without limits. When we allow the offspring body mass to evolve independently from adult body mass, it becomes smaller. However, when we take into account that larger individuals have larger offspring, both body masses evolve to larger values. These trends result from the allometric scaling of mortality and can be kept in limits by trade-offs other than those included in our model.

Geographic parthenogenesis in a consumer-resource model for sexual reproduction.

The phenomenon of geographic parthenogenesis is closely tied to the question of why sexual reproduction is the dominant mode of reproduction in animals and plants. Geographic parthenogenesis describes the fact that many species reproduce asexually at the boundaries of their range. We present a mathematical model that derives the dominance of sexuals at the center and the dominance of asexuals at the boundary of a species' range from exactly the same mechanism. Our model is based on a set of resources that regrow slowly and that can be consumed only by those individuals that have a suitable genotype. Genotype is implemented by a multilocus model with two alleles at each locus, and with free recombination during production of sexual offspring. The model is tailored to seasonal species with intermittent mixis and low survival of offspring, such as Daphnia and aphids. Several patches of resources are arranged in a row, with a gradient of those parameters that typically vary through the range of species. By letting sexually and asexually reproducing populations compete, we obtain the typical patterns of geographic parthenogenesis.