An outbreak of betacoronavirus severe acute respiratory syndrome (SARS)-CoV-2 began in Wuhan, China in December 2019. COVID-19, the disease associated with SARS-CoV-2 infection, rapidly spread to produce a global pandemic. We report development of a rapid (<40 min), easy-to-implement and accurate CRISPR-Cas12-based lateral flow assay for detection of SARS-CoV-2 from respiratory swab RNA extracts. We validated our method using contrived reference samples and clinical samples from patients in the United States, including 36 patients with COVID-19 infection and 42 patients with other viral respiratory infections. Our CRISPR-based DETECTR assay provides a visual and faster alternative to the US Centers for Disease Control and Prevention SARS-CoV-2real-time RT-PCR assay, with 95% positive predictive agreement and 100% negative predictive agreement.

Telomeres of eukaryotic chromosomes contain many tandem repeats of a G-rich sequence (for example, TTAGGG in vertebrates). In most normal human cells, telomeres shorten with each cell division, and it is proposed that this limits the number of times these cells can replicate. Telomeres may be maintained in germline cells, and in many immortalized cells and cancers, by the telomerase holoenzyme (first discovered in the ciliate Tetrahymena), which uses an RNA subunit as template for synthesis of telomeric DNA by the reverse transcriptase catalytic subunit. Some immortalized human cell lines and some tumours maintain their telomeres in the absence of any detectable telomerase activity by a mechanism referred to as alternative lengthening of telomeres (ALT). Here we show that DNA sequences are copied from telomere to telomere in an immortalized human ALT cell line, indicating that ALT occurs by means of homologous recombination and copy switching.

Summary The displacement loop (D-loop) is the DNA strand invasion product formed during homologous recombination. Disruption of nascent D-loops represents a mechanism of anti-recombination. During Synthesis-Dependent Strand Annealing D-loop disruption after extension of the invading strand is an integral step of the pathway and ensures a non-crossover outcome. The proteins implicated in D-loop disruption are DNA motor proteins/helicases acting by migrating DNA junctions. Here we report an unanticipated mechanism of D-loop dissolution mediated by DNA topoisomerase 3 (Top3) and dependent on its catalytic activity. D-loop dissolution catalyzed by yeast Top3 is highly specific for yeast Rad51/Rad54-mediated D-loops, whereas protein-free D-loops or D-loop mediated by bacterial RecA protein or human RAD51/RAD54 resist dissolution. Also the human Topoisomerase IIIα-RMI1–RMI2 complex is capable of dissolving D-loops. Consistent with genetic data, we suggest that the extreme growth defect and hyper-recombination phenotype of Top3-deficient yeast cells is in part a result of unprocessed D-loops.

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Carcinogenesis is a multistage process that has been characterized both by the activation of cellular oncogenes and by the loss of function of tumor suppressor genes. Colorectal cancer has been associated with the activation of ras oncogenes and with the deletion of multiple chromosomal regions including chromosomes 5q, 17p, and 18q. Such chromosome loss is often suggestive of the deletion or loss of function of tumor suppressor genes. The candidate tumor suppressor genes from these regions are, respectively, MCC and/or APC, p53, and DCC. In order to further our understanding of the molecular and genetic mechanisms involved in tumor progression and, thereby, of normal cell growth, it is important to determine whether defects in one or more of these loci contribute functionally in the progression to malignancy in colorectal cancer and whether correction of any of these defects restores normal growth control in vitro and in vivo. To address this question, we have utilized the technique of microcell-mediated chromosome transfer to introduce normal human chromosomes 5, 17, and 18 individually into recipient colorectal cancer cells. Additionally, chromosome 15 was introduced into SW480 cells as an irrelevant control chromosome. While the introduction of chromosome 17 into the tumorigenic colorectal cell line SW480 yielded no viable clones, cell lines were established after the introduction of chromosomes 15, 5, and 18. Hybrids containing chromosome 18 are morphologically similar to the parental line, whereas those containing chromosome 5 are morphologically distinct from the parental cell line, being small, polygonal, and tightly packed. SW480-chromosome 5 hybrids are strongly suppressed for tumorigenicity, while SW480-chromosome 18 hybrids produce slowly growing tumors in some of the animals injected. Hybrids containing the introduced chromosome 18 but was significantly reduced in several of the tumor reconstitute cell lines. Introduction of chromosome 5 had little to no effect on responsiveness, whereas transfer ot chromosome 18 restored responsiveness to some degree. Our findings indicate that while multiple defects in tumor suppressor genes seem to be required for progression to the malignant state in colorectal cancer, correction of only a single defect can have significant effects in vivo and/or in vitro.