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PURPOSE A substantial proportion of patients enrolled on ACOSOG Z0011 received protocol-deviant radiation treatment. It is currently unknown whether these deviations involved the use of more extensive fields in patients at higher nomogram-predicted risk. METHODS We used the M.D. Anderson (MDA) and Memorial Sloan-Kettering (MSK) nomograms to estimate risk of additional positive axillary nodes using surgical pathology information. In the control arm, we compared axillary dissection (AD) findings to nomogram-predicted estimates for validation. We used logistic regression to evaluate whether nomogram-estimated higher risk of nodal involvement was associated with high tangent (HT) or supraclavicular (SCV) radiation fields for patients with known radiation field design. RESULTS 552/856 (64.5%) had complete details for the MDA nomogram. Mean MDA risk estimate in both treatment arms was 23.8%. Estimated risk for patients on the AD arm with positive nodes was 25.9%. Higher risk estimate was associated with additional positive nodes in the AD arm (OR 1.04, 95% CI 1.02-1.06, p less then 0.0001). We observed significant association with higher MDA nomogram-estimated risk and SCV radiation (OR 1.07, 95% CI 1.04-1.10, p less then 0.0001) but not HT (OR 0.99, 95% CI 0.96-1.02, p = 0.52) The MSK nomogram had similar associations. CONCLUSION MDA and MSK nomogram risk estimates were associated with lymph node risk in ACOSOG Z0011. Radiation oncologists' use of differing radiation fields were associated with treating higher risk patients. ClinicalTrials.gov id NCT00003854.Here, for the first time, we have investigated the hipBAXn toxin-antitoxin (TA) module from entomopathogenic bacterium Xenorhabdus nematophila. It is a type II TA module that consists of HipAXn toxin and HipBXn antitoxin protein and located in the complementary strand of chromosome under XNC1_operon 0810 locus tag. For functional analysis, hipAXn toxin, hipBXn antitoxin, and an operon having both genes were cloned in pBAD/His C vector and transformed in Escherichia coli cells. The expression profiles and endogenous toxicity assay were performed in these cells. To determine the active amino acid residues responsible for the toxicity of HipAXn toxin, site-directed mutagenesis (SDM) was performed. SDM results showed that amino acid residues S149, D306, and D329 in HipAXn toxin protein were significantly essential for its toxicity. For transcriptional analysis, the 157 bp upstream region of the hipBAXn TA module was identified as a promoter with bioinformatics tools. Further, the LacZ reporter construct with promoter region was prepared and LacZ assays as well as reverse transcriptase-polymerase chain reaction (RT-PCR) analysis was performed under different stress conditions. Electrophoretic mobility shift assay (EMSA) was also performed with recombinant HipAXn toxin, HipBXn antitoxin protein, and 157 bp promoter region. Results showed that the hipBAXn TA module is a well-regulated system in which the upregulation of gene expression was also found compulsive in different SOS conditions. KEY POINTS •Functional characterization of hipBA Xn TA module from Xenorhabdus nematophila. •hipBA Xn TA module is a functional type II TA module. •Transcriptional characterization of hipBA Xn TA module. •hipBA Xn TA module is a well regulated TA module. Graphical abstract.Microbe-based decontamination of phenol-polluted environments has significant advantages over physical and chemical approaches by being relatively cheaper and ensuring complete phenol degradation. There is a need to search for commercially prospective bacterial strains that are resistant to phenol and other co-pollutants, e.g. oil hydrocarbons, in contaminated environments, and able to carry out efficient phenol biodegradation at a variable range of concentrations. This research characterizes the phenol-biodegrading ability of a new actinobacteria strain isolated from a lubricant-contaminated soil environment. Phenotypic and phylogenetic analyses showed that the novel strain UCM Ac-603 belonged to the species Rhodococcus aetherivorans, and phenol degrading ability was quantitatively characterized for the first time. selleckchem R. aetherivorans UCM Ac-603 tolerated and assimilated phenol (100% of supplied concentration) and various hydrocarbons (56.2-94.4%) as sole carbon sources. Additional nutrient supplementation was environments.Targeted gene mutation by allelic replacement is important for functional genomic analysis and metabolic engineering. However, it is challenging in mutating the essential genes with the traditional method by using a selection marker, since the first step of essential gene knockout will result in a lethal phenotype. Here, we developed a two-end selection marker (Two-ESM) method for site-directed mutation of essential genes in Saccharomyces cerevisiae with the aid of the CRISPR/Cas9 system. With this method, single and double mutations of the essential gene ERG20 (encoding farnesyl diphosphate synthase) in S. cerevisiae were successfully constructed with high efficiencies of 100%. In addition, the Two-ESM method significantly improved the mutation efficiency and simplified the genetic manipulation procedure compared with traditional methods. The genome integration and mutation efficiencies were further improved by dynamic regulation of mutant gene expression and optimization of the integration modules. This Two-ESM method will facilitate the construction of genomic mutations of essential genes for functional genomic analysis and metabolic flux regulation in yeasts. KEY POINTS • A Two-ESM strategy achieves mutations of essential genes with high efficiency of 100%. • The optimized three-module method improves the integration efficiency by more than three times. • This method will facilitate the functional genomic analysis and metabolic flux regulation.Monascus is a filamentous fungus that produces several secondary metabolites. Here, we investigated the effects of the global regulator LaeA on the synthesis of pigments and monacolin K in Monascus purpureus with spectrophotometer and HPLC methods. The LaeA gene was isolated from M. purpureus M1 to create an overexpression construct. An LaeA-overexpressing strain L3 was with 48.6% higher monacolin K production than the M1 strain. The L3 strain also produced higher Monascus pigments than the M1 strain. SEM showed that LaeA overexpression resulted in altered mycelial morphology. Compared with the M1 strain, the L3 strain expressed higher levels of monacolin K synthesis-related genes mokA, mokB, mokE, and mokH. Overall, these results suggest that LaeA plays a role in regulating the production of secondary metabolites and mycelial growth in Monascus. This study provides important insights into the mechanisms underlying the effects of the LaeA gene on the secondary metabolites of M. purpureus.
Website: https://www.selleckchem.com/products/sodium-phenylbutyrate.html
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