RNA vaccines have become a key tool in moving forward through the challenges raised both in the current pandemic and in numerous other public health and medical challenges. With the rollout of vaccines for COVID-19, these synthetic mRNAs have become broadly distributed RNA species in numerous human populations. Despite their ubiquity, sequences are not always available for such RNAs. Standard methods facilitate such sequencing. In this note, we provide experimental sequence information for the RNA components of the initial Moderna (https://pubmed.ncbi.nlm.nih.gov/32756549/) and Pfizer/BioNTech (https://pubmed.ncbi.nlm.nih.gov/33301246/) COVID-19 vaccines, allowing a working assembly of the former and a confirmation of previously reported sequence information for the latter RNA. Sharing of sequence information for broadly used therapeutics has the benefit of allowing any researchers or clinicians using sequencing approaches to rapidly identify such sequences as therapeutic-derived rather than host or infectious in origin. For this work, RNAs were obtained as discards from the small portions of vaccine doses that remained in vials after immunization; such portions would have been required to be otherwise discarded and were analyzed under FDA authorization for research use. To obtain the small amounts of RNA needed for characterization, vaccine remnants were phenol-chloroform extracted using TRIzol Reagent (Invitrogen), with intactness assessed by Agilent 2100 Bioanalyzer before and after extraction. Although our analysis mainly focused on RNAs obtained as soon as possible following discard, we also analyzed samples which had been refrigerated (~4 ℃) for up to 42 days with and without the addition of EDTA. Interestingly a substantial fraction of the RNA remained intact in these preparations. We note that the formulation of the vaccines includes numerous key chemical components which are quite possibly unstable under these conditions-- so these data certainly do not suggest that the vaccine as a biological agent is stable. But it is of interest that chemical stability of RNA itself is not sufficient to preclude eventual development of vaccines with a much less involved cold-chain storage and transportation. For further analysis, the initial RNAs were fragmented by heating to 94℃, primed with a random hexamer-tailed adaptor, amplified through a template-switch protocol (Takara SMARTerer Stranded RNA-seq kit), and sequenced using a MiSeq instrument (Illumina) with paired end 78-per end sequencing. As a reference material in specific assays, we included RNA of known concentration and sequence (from bacteriophage MS2). From these data, we obtained partial information on strandedness and a set of segments that could be used for assembly. This was particularly useful for the Moderna vaccine, for which the original vaccine RNA sequence was not available at the time our study was carried out. Contigs encoding full-length spikes were assembled from the Moderna and Pfizer datasets. The Pfizer/BioNTech data [Figure 1] verified the reported sequence for that vaccine (https://berthub.eu/articles/posts/reverse-engineering-source-code-of-the-biontech-pfizer-vaccine/), while the Moderna sequence [Figure 2] could not be checked against a published reference. RNA preparations lacking dsRNA are desirable in generating vaccine formulations as these will minimize an otherwise dramatic biological (and nonspecific) response that vertebrates have to double stranded character in RNA (https://www.nature.com/articles/nrd.2017.243). In the sequence data that we analyzed, we found that the vast majority of reads were from the expected sense strand. In addition, the minority of antisense reads appeared different from sense reads in lacking the characteristic extensions expected from the template switching protocol. Examining only the reads with an evident template switch (as an indicator for strand-of-origin), we observed that both vaccines overwhelmingly yielded sense reads (>99.99%). Independent sequencing assays and other experimental measurements are ongoing and will be needed to determine whether this template-switched sense read fraction in the SmarterSeq protocol indeed represents the actual dsRNA content in the original material. This work provides an initial assessment of two RNAs that are now a part of the human ecosystem and that are likely to appear in numerous other high throughput RNA-seq studies in which a fraction of the individuals may have previously been vaccinated. ProtoAcknowledgements: Thanks to our colleagues for help and suggestions (Nimit Jain, Emily Greenwald, Lamia Wahba, William Wang, Amisha Kumar, Sameer Sundrani, David Lipman, Bijoyita Roy). Figure 1: Spike-encoding contig assembled from BioNTech/Pfizer BNT-162b2 vaccine. Although the full coding region is included, the nature of the methodology used for sequencing and assembly is such that the assembled contig could lack some sequence from the ends of the RNA. Within the assembled sequence, this hypothetical sequence shows a perfect match to the corresponding sequence from documents available online derived from manufacturer communications with the World Health Organization [as reported by https://berthub.eu/articles/posts/reverse-engineering-source-code-of-the-biontech-pfizer-vaccine/]. The 5’ end for the assembly matches the start site noted in these documents, while the read-based assembly lacks an interrupted polyA tail (A30(GCATATGACT)A70) that is expected to be present in the mRNA.
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An #OSINT Framework to perform various recon techniques on Companies, People, Phone Number, Bitcoin Addresses, etc., aggregate all the raw data, and give data in multiple formats.
Investigate malicious Windows logon by visualizing and analyzing Windows event log
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I created this repository to keep my learning, notes, and code in one place for various tools in DevOps. Now, it's helping thousands of learners, practitioners, and professionals every day in their DevOps journey.
- docker
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GitHub action to set up PHP with extensions, php.ini configuration, coverage drivers, and various tools.
- php
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ANDROID. ChipsLayoutManager (SpanLayoutManager, FlowLayoutManager). A custom layout manager for RecyclerView which mimicric TextView span behaviour, flow layouts behaviour with support of amazing recyclerView features
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Multifarious Scrapy examples. Spiders for alexa / amazon / douban / douyu / github / linkedin etc.
Intermediate Graphics Library (IGL) is a cross-platform library that commands the GPU. It provides a single low-level cross-platform interface on top of various graphics APIs (e.g. OpenGL, Metal and Vulkan).
SteamKit2 is a .NET library designed to interoperate with Valve's Steam network. It aims to provide a simple, yet extensible, interface to perform various actions on the network.
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Provides integration for Doctrine with various Symfony components
:whale: A curated list of delicious docker recipes 🇺🇦🇮🇱 (Let's Fight Against Dictatorship)
Article publishing platform that automatically distributes your articles to various media channels
An open source version of a game engine developed by Croteam for the classic Serious Sam games.
A Webpack plugin to enable "Fast Refresh" (also previously known as Hot Reloading) for React components.
- webpack-plugin
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- react
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the LIBpcap interface to various kernel packet capture mechanism
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Mod manager for various PC games. Discord Server: https://discord.gg/ewUVAqyrQX if you would like to be more involved
BrowserFS is an in-browser filesystem that emulates the Node JS filesystem API and supports storing and retrieving files from various backends.
WarpDrive is the lightweight data library for ambitious web apps — universal, typed, reactive, and ready to scale.
- ember
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Web application for archival and reading of manga/doujinshi. Lightweight and Docker-ready for NAS/servers.
- perl
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第三方支付对接全能支付Java开发工具包.优雅的轻量级支付模块集成支付对接支付整合（微信,支付宝,银联,友店,富友,跨境支付paypal,payoneer(P卡派安盈)易极付）app,扫码,网页刷脸付刷卡付条码付转账服务商模式,微信分账,微信合单支付、支持多种支付类型多支付账户，支付与业务完全剥离，简单几行代码即可实现支付，简单快速完成支付模块的开发，可轻松嵌入到任何系统里 目前仅是一个开发工具包（即SDK），只提供简单Web实现，建议使用maven或gradle引用本项目即可使用本SDK提供的各种支付相关的功能
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Various translations of OSTEP can be found here. Help the cause and contribute!
ReportGenerator converts coverage reports generated by coverlet, OpenCover, dotCover, Visual Studio, NCover, Cobertura, JaCoCo, Clover, gcov or lcov into human readable reports in various formats.
- code-coverage
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A collection of my book notes on various subjects, mainly computer science
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Boilerplate configurations for various web servers.