The transcription activator-like effectors (TALEs) and the RNA-guided clustered regularly interspaced short palindromic repeat (CRISPR) associated protein (Cas9) utlilize distinct molecular ...mechanisms in targeting site recognition. The two proteins can be modified to carry additional functional domains to regulate expression of genomic loci in mammalian cells. In this study, we have compared the two systems in activation and suppression of the Oct4 and Nanog loci by targeting their enhancers. Although both are able to efficiently activate the luciferase reporters, the CRISPR/dCas9 system is much less potent in activating the endogenous loci and in the application of reprogramming somatic cells to iPS cells. Nevertheless, repression by CRISPR/dCas9 is comparable to or even better than TALE repressors. We demonstrated that dCas9 protein binding results in significant physical interference to binding of native transcription factors at enhancer, less efficient active histone markers induction or recruitment of activating complexes in gene activation. This study thus highlighted the merits and drawbacks of transcription regulation by each system. A combined approach of TALEs and CRISPR/dCas9 should provide an optimized solution to regulate genomic loci and to study genetic elements such as enhancers in biological processes including somatic cell reprogramming and guided differentiation.
Core stabilization has been utilized for rehabilitation and prevention of lower limb musculoskeletal injuries. Previous studies showed that activation of the abdominal core muscles enhanced the hip ...muscle activity in hip extension and abduction exercises. However, the lack of the direct measurement and quantification of the activation level of the abdominal core muscles during the execution of the hip exercises affect the level of evidence to substantiate the proposed application of core exercises to promote training and rehabilitation outcome of the hip region. The aim of the present study was to examine the effects of abdominal core activation, which is monitored directly by surface electromyography (EMG), on hip muscle activation while performing different hip exercises, and to explore whether participant characteristics such as gender, physical activity level and contractile properties of muscles, which is assessed by tensiomyography (TMG), have confounding effect to the activation of hip muscles in enhanced core condition.
Surface EMG of bilateral internal obliques (IO), upper gluteus maximus (UGMax), lower gluteus maximus (LGMax), gluteus medius (GMed) and biceps femoris (BF) of dominant leg was recorded in 20 young healthy subjects while performing 3 hip exercises: Clam, side-lying hip abduction (HABD), and prone hip extension (PHE) in 2 conditions: natural core activation (NC) and enhanced core activation (CO). EMG signals normalized to percentage of maximal voluntary isometric contraction (%MVIC) were compared between two core conditions with the threshold of the enhanced abdominal core condition defined as >20%MVIC of IO.
Enhanced abdominal core activation has significantly promoted the activation level of GMed in all phases of clam exercise (P < 0.05), and UGMax in all phases of PHE exercise (P < 0.05), LGMax in eccentric phases of all 3 exercises (P < 0.05), and BF in all phases of all 3 exercises except the eccentric phase of PHE exercise (P < 0.05). The %MVIC of UGMax was significantly higher than that of LGMax in all phases of clam and HABD exercises under both CO and NC conditions (P < 0.001) while the %MVIC of LGMax was significantly higher than UGMax in concentric phase of PHE exercise under NC condition (P = 0.003). Gender, physical activity level and TMG parameters were not major covariates to activation of hip muscles under enhanced core condition.
Abdominal core activation enhances the hip muscles recruitment in Clam, HABD and PHE exercises, and this enhancement is correlated with higher physical activity and stiffer hip muscle. Our results suggest the potential application of abdominal core activation for lower limb rehabilitation since the increased activation of target hip muscles may enhance the therapeutic effects of hip strengthening exercises.
Recent studies have suggested that single-stranded DNA (ssDNA) library preparation can enrich short DNA species from the plasma of healthy individuals, cancer patients, and transplant recipients. ...Based on previous observations that fetal DNA molecules in the maternal plasma are shorter than maternal DNA molecules, ssDNA library preparation may potentially enrich fetal DNA and provide substantial improvement in noninvasive prenatal testing.
We tested this hypothesis by comparing the maternal plasma DNA sequencing results using 2 types of ssDNA library preparation methods and a standard double-stranded DNA (dsDNA) library method using samples from first- and third-trimester pregnancies. We also evaluated the performance of ssDNA and dsDNA library methods in the noninvasive prenatal detection of trisomy 21 from maternal plasma.
Short DNA species were significantly enriched in ssDNA libraries. However, contrary to previous speculation, no significant enrichment was observed in the overall fetal fraction in maternal plasma collected in the first trimester. Our use of an ssDNA library did not reduce the variation in chromosomal representation when compared with a standard dsDNA library in the first-trimester plasma samples. ssDNA libraries also showed inferior performance in the noninvasive prenatal detection of trisomy 21 from maternal plasma. Detailed fetal fraction analysis using size-fractionated Y chromosome sequences and fetal-specific single-nucleotide polymorphisms (SNPs) revealed an unexpected finding that short maternal DNA was preferentially enriched over short fetal DNA in an ssDNA library irrespective of GC content.
Our findings have shown that ssDNA library preparation preferentially enriches short maternally derived DNA in maternal plasma.
Sustainable consumption and production is a critical issue in the chemical industry due to increasing public concerns on environmental and safety issues. Organizations are urged to improve the ...quality of chemical products while minimizing the environmental impacts during production. In current practice, chemists and formulators have to determine both the ingredients to be used and the machine parameter settings during product development and production. Without appropriate operations strategies for managing sustainable consumption and production, a significant portion of the ingredients, toxic materials and pollutants are wasted or emitted during the trial-and-error processes when developing chemical products. In addition, inappropriate machine parameter settings, such as blending speed and blending temperature, result in inefficient energy use. Motivated by these issues, this paper describes a recursive operations strategy (ROS) model for achieving sustainable consumption and production in the chemical industry. The ROS model first identifies the business strategy, and then defines operations strategies by assessing the competitive priorities and policies with the use of artificial intelligence, including case-based reasoning and fuzzy logic, so as to manage the operations functions. The effectiveness of the model is verified by means of a case study. The results indicate that the model can provide direct guidelines for the users to develop products based on previously developed products. By so doing, the number of trials for testing various ingredient formulae can be reduced, minimizing the ingredient waste. The proposed model is also capable of achieving continuous improvement and determining the optimal production process conditions for avoiding unnecessary energy consumption.
•To support the sustainable chemical product development and production.•To design policies in the selection of chemical ingredients.•To define operations strategy for appropriate production process conditions.•To minimize chemical waste by reducing the number of trials in product formulation.•To avoid unnecessary energy consumption by adopting the optimal process parameters.
The specific detection of a minor population of mutant DNA molecules requires methods of high specificity and sensitivity. While the single-allele base extension reaction (SABER) was shown to be ...useful for the detection of certain beta-thalassemia mutations, we encountered problems with false positivity during development of SABER for the noninvasive prenatal diagnosis of the hemoglobin E (HbE) disease. Systematic optimization resulted in an alternative protocol, the allele-specific base extension reaction (ASBER).
An artificial model was established by mixing genomic DNA of HbE carriers and normal individuals. Effects of terminator concentration and annealing temperature on the nonspecificity of SABER were then studied. The use of a single relevant terminator and the other 3 types of dideoxynucleotide as competing terminators were also compared in the development of the ASBER protocol. Thirteen cases of HbE-susceptible pregnancies were tested to compare the SABER and the ASBER protocols.
Decreasing the single relevant terminator concentration and increasing the annealing temperature in SABER were found to improve specificity. The use of the other 3 types of dideoxynucleotide as competing terminators was shown to offer better detection sensitivity than a single terminator in ASBER. Genotyping results were all correctly determined by ASBER, except one false-negative detection (sensitivity: 80%, specificity: 100%).
An alternative mass spectrometry-based protocol for noninvasive prenatal diagnosis, ASBER, has been successfully developed to allow the detection of a minor DNA population with a point mutation.
Abstract
Interplay between EBV infection and acquired genetic alterations during nasopharyngeal carcinoma (NPC) development remains vague. Here we report a comprehensive genomic analysis of 70 NPCs, ...combining whole-genome sequencing (WGS) of microdissected tumor cells with EBV oncogene expression to reveal multiple aspects of cellular-viral co-operation in tumorigenesis. Genomic aberrations along with EBV-encoded LMP1 expression underpin constitutive NF-κB activation in 90% of NPCs. A similar spectrum of somatic aberrations and viral gene expression undermine innate immunity in 79% of cases and adaptive immunity in 47% of cases; mechanisms by which NPC may evade immune surveillance despite its pro-inflammatory phenotype. Additionally, genomic changes impairing
TGFBR2
promote oncogenesis and stabilize EBV infection in tumor cells. Fine-mapping of
CDKN2A/CDKN2B
deletion breakpoints reveals homozygous
MTAP
deletions in 32-34% of NPCs that confer marked sensitivity to MAT2A inhibition. Our work concludes that NPC is a homogeneously NF-κB-driven and immune-protected, yet potentially druggable, cancer.
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