Background
The recent emergence of SARS-CoV-2 lead to a current pandemic of unprecedented scale. Though diagnostic tests are fundamental to the ability to detect and respond, overwhelmed healthcare systems are already experiencing shortages of reagents associated with this test, calling for a lean immediately-applicable protocol.
Methods
RNA extracts of positive samples were tested for the presence of SARS-CoV-2 using RT-qPCR, alone or in pools of different sizes (2-, 4-, 8- ,16-, 32- and 64-sample pools) with negative samples. Transport media of additional 3 positive samples were also tested when mixed with transport media of negative samples in pools of 8.
Results
A single positive sample can be detected in pools of up to 32 samples, using the standard kits and protocols, with an estimated false negative rate of 10%. Detection of positive samples diluted in even up to 64 samples may also be attainable, though may require additional amplification cycles. Single positive samples can be detected when pooling either after or prior to RNA extraction.
Conclusions
As it uses the standard protocols, reagents and equipment, this pooling method can be applied immediately in current clinical testing laboratories. We hope that such implementation of a pool test for COVID-19 would allow expanding current screening capacities thereby enabling the expansion of detection in the community, as well as in close organic groups, such as hospital departments, army units, or factory shifts.
Regulation of gene expression is executed in many cases by RNA-binding proteins
(RBPs) that bind to mRNAs as well as to non-coding RNAs. RBPs recognize their
RNA target via specific binding sites on the RNA. Predicting the binding sites
of RBPs is known to be a major challenge. We present a new webserver, RBPmap,
freely accessible through the website http://rbpmap.technion.ac.il/ for accurate prediction and
mapping of RBP binding sites. RBPmap has been developed specifically for mapping
RBPs in human, mouse and Drosophila melanogaster genomes,
though it supports other organisms too. RBPmap enables the users to select
motifs from a large database of experimentally defined motifs. In addition,
users can provide any motif of interest, given as either a consensus or a PSSM.
The algorithm for mapping the motifs is based on a Weighted-Rank approach, which
considers the clustering propensity of the binding sites and the overall
tendency of regulatory regions to be conserved. In addition, RBPmap incorporates
a position-specific background model, designed uniquely for different genomic
regions, such as splice sites, 5’ and 3’ UTRs, non-coding RNA
and intergenic regions. RBPmap was tested on high-throughput RNA-binding
experiments and was proved to be highly accurate.
This article presents an overview of the SAM-T02 method for protein fold recognition and the UNDERTAKER program for ab initio predictions. The SAM-T02 server is an automatic method that uses two-track hidden Markov models (HMMS) to find and align template proteins from PDB to the target protein. The two-track HMMs use an amino acid alphabet and one of several different local structure alphabets. The UNDERTAKER program is a new fragment-packing program that can use short or long fragments and alignments to create protein conformations. The HMMs and fold-recognition alignments from the SAM-T02 method were used to generate the fragment and alignment libraries used by UNDERTAKER. We present results on a few selected targets for which this combined method worked particularly well: T0129, T0181, T0135, T0130, and T0139.
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