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The CRISPR/Cas9 system has unprecedentedly revolutionized genome-editing technology, which is being successfully applied virtually in all branches of biological sciences. Although much success has been attained in gene manipulation, still the majority of methods are laborious and non-integration-free, and require prolonged time for the expansion of mutant cell pools/clones, while fewer cells exhibit functional knockout efficiency. To overcome these obstacles, here, we describe an efficient, inexpensive, integration-free, and rapid one-step protocol for CRISPR/Cas9-assisted gene knockout in murine pluripotent stem cells (PSCs). Our protocol has streamlined both the liposome-based transfection system and screening strategy to work more efficiently with small numbers of PSCs (∼2.0 × 104 cells) and to minimize laborious steps of lentiviral packaging, transduction, and single-clone passaging. In our method, around 90% (CI = 95%, 79.5230%–100%) of PSC colonies harbored functional knockout in the context of protein expression. Therefore, the current protocol is technically feasible, time-saving, and highly efficient for genome editing in pluripotent stem cells.

The emergence of the COVID-19 Delta variant-of-concern (VOC), a novel variant of SARS-CoV-2, has threatened the total health systems throughout the world. This highly contagious strain is spreading at a higher exponential rate than any other variants of COVID-19 by infecting and subsequently killing hundreds of thousands of people globally. Among the most sensitive groups, pregnant women are at high risk of increased hospitalization, pneumonia, respiratory support, and admission to intensive care units during the Delta period. Pregnant people with gestational diabetes mellitus (GDM) are at increased chances of Delta VOC infection. GDM patients are nine and three times more likely to be infected by Delta VOC than those pregnant patients suffering from diabetes and cardiovascular diseases and hypertension, respectively. Additionally, they are more vulnerable to Delta VOC infection than wild-type and Alpha COVID-19 VOC ones. Thus, this review critically sheds light on the current scenario of the vulnerability of pregnant mothers, especially those with GDM, to Delta VOC infection.

Background: Previously, we demonstrated that a water, sanitation, handwashing, and nutritional intervention improved linear growth and was unexpectedly associated with shortened childhood telomere length (TL) (Lin et al., 2017). Here, we assessed the association between TL and growth. Methods: We measured relative TL in whole blood from 713 children. We reported differences between the 10th percentile and 90th percentile of TL or change in TL distribution using generalized additive models, adjusted for potential confounders. Results: In cross-sectional analyses, long TL was associated with a higher length-for-age Z score at age 1 year (0.23 SD adjusted difference in length-for-age Z score [95% CI 0.05, 0.42; FDR-corrected p-value = 0.01]). TL was not associated with other outcomes. Conclusions: Consistent with the metabolic telomere attrition hypothesis, our previous trial findings support an adaptive role for telomere attrition, whereby active TL regulation is employed as a strategy to address ‘emergency states’ with increased energy requirements such as rapid growth during the first year of life. Although short periods of active telomere attrition may be essential to promote growth, this study suggests that a longer overall initial TL setting in the first 2 years of life could signal increased resilience against future telomere erosion events and healthy growth trajectories. Funding: Funded by the Bill and Melinda Gates Foundation. Clinical trial number: NCT01590095