8 skills found
NAalytics / Assemblies Of Putative SARS CoV2 Spike Encoding MRNA Sequences For Vaccines BNT 162b2 And MRNA 1273RNA 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.
johnkil / Android ProgressFragmentImplementation of the fragment with the ability to display indeterminate progress indicator when you are waiting for the initial data.
kevinGodell / Mp4fragParser that works with ffmpeg to read piped data and fragment mp4 into an initialization segment and media segments. It can also get the codec info and generate an fmp4 HLS m3u8 playlist.
dilpreet2028 / Fragmenter[Android Library] Remove Boilerplate code for initializing fragments
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Ram15144 / Guess Game Android StudioA strategy game called Guess Four in which each of two players sets up a secret sequence of four decimal digits. The two players take turns guessing the sequence of digits of their opponent. A player responds to an opponents guess by specifying the number of digits that were successfully guessed in the correct position and the number of digits that were successfully guessed but in the wrong positions. Thus, if a players chosen number is 2017 and the opponent guesses 1089, the opponent would be told that one digit was guessed correctly in the correct position (i.e., the 0), and another digit was guessed in the wrong position, (i.e., the 1). The implementation involves two background threads that play against each other while also updating the user interface with their moves. The first thread to correctly guess the opposing thread’s number wins the game. The UI thread is responsible for creating and starting the two worker threads and for maintaining and updating the display. Each worker thread will take turns taking the following actions: 1. Waiting for a short time (1-2 seconds) in order for a human viewer to take note of the previous move on the display. 2. Figuring out the next guess of this thread. 3. Communicating this guess to the opponent thread. 4. Waiting for a response from the opponent thread. 5. Communicating both the guess and the response from the opposite thread to the UI thread. While carrying out the above steps each worker thread is able to respond to guesses from the opponent thread. Whenever a guess from the opponent thread is received, a worker thread responds by communicating the number of correctly positioned and incorrectly positioned digits contained in the guess to the opponent thread. Furthermore, the game must proceeds in lockstep between the two threads. A thread is not allowed to make two consecutive guesses without handling an intervening guess from the opponent thread. The UI thread is specifically responsible for the following functionality: 1. Showing the two initial numbers chosen by the worker threads. 2. Receiving notifications of guesses and their outcomes by the worker threads. 3. Displaying the guesses and responses of each worker thread in an appropriate format as each worker thread communicates this information to the UI thead. 4. Displaying a button to start the game. Pressing this button while a game is in progress will void the current game and start a new game from scratch. 5. Checking on the status of the game, by determining whether one thread has won or the game needs to continue. 6. Halting the game after each thread has made 20 guesses without identifying the opponent’s number. 7. Signaling the two worker threads that the game is over; the two threads stops their execution as a result of this action. Displaying the outcome of the game in the UI. Implementation: 1. Handlers are used to implement the communication between the three threads involved. Each thread has a handler, a job queue and a looper. 2. Both runnables and messages are included in the job queue of the worker threads. 3. The two worker threads uses different strategies for winning the game. 4. The game is played at such a speed that a human user can clearly see and understand the move of each thread. 5. The app’s display has two layouts or fragments, one for each worker thread, lying side-by-side and occupying the entire screen width. Each side displays the secret number of the corresponding thread at the top and a scrollable list of the thread’s guesses and the opponent thread’s responses in the remainder of the vertical space. 7. The two worker threads will use a random number generator to create their secret number. These numbers are communicated to the UI thread, which will immediately display both numbers.
Lewenhaupt / CtxEasily initialize AGENTS.md, GEMINI.md, CLAUDE.md, etc. files from tagged fragment.md files
gunhansancar / FragmentPagerExampleThis is a sample application to show how to use fragment pager by utilizing the factory pattern to initialize fragment instances.
