Himanshu Mishra’s Post

What is CRISPR-Cas9? CRISPR-Cas9 is a genome editing technology used to change parts of the genome by removing, adding, or altering sections of DNA. It uses a pre-designed RNA sequence to guide the Cas9 enzyme to the location where the genome needs to be altered. The Cas9 enzyme then cleaves the DNA strands, and the cell's natural repair mechanism joins the two ends together by removing, adding, or replacing certain sections.

DNA structure and Chromosome composition DNA is double stranded, and the strands are antiparallel because they run in opposite directions. Explanation: Each DNA molecule has two strands of nucleotides. Each strand has sugar phosphate backbone, but the orientation of the sugar molecule is opposite in the two strands. Both of the strands of DNA double helix can grow in 5' to 3' direction, but they grow in opposite directions due to opposite orientation of the sugar molecule in them. The antiparallel orientation allows for the base pairs to compliment one another. Antiparallel DNA is also more structurally stable than parallel DNA. The antiparallel orientation of DNA has important implications for DNA replication, as at the replication fork one strand allows steady replication, thereby known as leading strand while the other becomes lagging strand. A histone is a protein that provides structural support to a chromosome. In order for very long DNA molecules to fit into the cell nucleus, they wrap around complexes of histone proteins, giving the chromosome a more compact shape. Some variants of histones are associated with the regulation of gene expression. Problems: What is meant by the description "antiparallel" regarding the two strands that make up the DNA double helix? A\) The 5' to 3' direction of one strand runs counter to the 5' to 3' direction of the other strand. B\) One strand contains only purines and the other contains only pyrimidines. C\) Base pairings create unequal spacing between the two DNA strands. D\) The double helix structure of DNA creates nonparallel strands. Eukaryotic chromosomes are composed of which of the following macromolecules? A\) DNA only B\) DNA and proteins C\) DNA and RNA D\) DNA and phospholipids Youtube video: https://lnkd.in/dmhsTuwD \#nikolays_genetics_lessons

The research highlights a significant advance in the field of genetic engineering by focusing on IS110 insertion sequences. These sequences express a structured non-coding RNA, termed a bridge RNA, which specifically binds to the encoded recombinase. The bridge RNA features two internal loops that base-pair with the target DNA and the donor DNA, enabling precise sequence-specific recombination. This system’s modularity allows for customizable DNA insertion, excision, and inversion, offering a versatile tool for genome manipulation. This innovation holds promise for advancing genome design and enhancing genetic diversity through targeted genomic rearrangements. Read more Durrant, M.G., Perry, N.T., Pai, J.J. et al. Bridge RNAs direct programmable recombination of target and donor DNA. Nature 630, 984–993 (2024). https://lnkd.in/gZ_GM3i5

Whole genome amplification (WGA) is a key step in DNA sequencing from low-input DNA samples, such as forensics materials, cell-free DNA from blood, and spent media from cell culture. Commonly used methods, such as DOP-PCR, MDA, MALBAC, and LIANTI, all have drawbacks such as amplification bias, poor uniformity, errors and artifacts, low genome coverage, inability to address all variant classes, low accuracy, poor reproducibility, and/or complex protocols that are difficult to automate or scale. Primary template-directed amplification (PTA) is a novel, isothermal WGA method that reproducibly captures >95% of the genomes of single cells, in a controlled and more uniform and accurate manner than existing approaches. This improves variant calling sensitivity and specificity, lowers sequencing costs, and facilitates bioinformatic analysis. - PTA enables superior genomic coverage versus other single-cell whole genome methods and is comparable to bulk sequencing - PTA enables superior single nucleotide variant (SNV) detection versus other single-cell whole genome methods and is comparable to bulk sequencing - PTA enables superior copy number variation (CNV) detection versus other single-cell whole genome methods and is comparable to bulk sequencing Get more from your DNA sequencing – request a quote for our ResolveDNA WGA Kits today! ⤵ https://lnkd.in/g-XptHu7

RNA programmable recombinases This pre-print (not yet peer reviewed) from Patrick Hsu's lab, describes a group of designer recombinases. Developed from a type of mobile genetic element that encodes its own “bridge RNA” to direct its integration, the paper describes how both the RNA bridge/guide and the target site can be reprogrammed to direct insertions, excision and inversion. A *huge* thank you to the two first authors, Matthew Durrant and Nicholas Perry, for their excellent webinar today talking us through the paper. I can’t wait to see if it will work in mammalian cells as well as E.coli! #GenomeEngineering #recombinase #IS110

A comparative analysis of 404 Monkeypox virus (MPXV) genomes has revealed significant changes in microsatellite abundance and density, particularly within Clades I, IIa, and IIb. Key findings include: 🔬 Unique microsatellite motifs for each clade, with twenty-six conserved loci specific to MPXV, highlighting their potential as molecular markers in diagnostics. 🔬 Identification of nine genes in the MPXV genome featuring ten variable hotspot microsatellite regions, associated with surface protein synthesis and host control. 🔬 Notable variations in the gene OPG153, especially at the SSR locus ‘(ATC)n’, which showed pronounced changes across lineages over time and plays a crucial role in virus pathogenesis within the host cell. These findings enhance our understanding of MPXV's unique molecular profile and provide valuable insights into its potential pathogenic and evolutionary implications. #Monkeypox #MPXV #Genomics #PublicHealth #Virology #InfectiousDiseases #MolecularBiology #Diagnostics https://lnkd.in/gJfCsXBH

Mechanisms for HNH-mediated target DNA cleavage in type I #CRISPR-Cas systems:- •The metagenome-derived type I-E and type I-F variant CRISPR-associated complex for antiviral defense (Cascade) complexes, fused with HNH domains, precisely cleave target DNA, representing recently identified genome editing tools. •However, the underlying working mechanisms remain unknown. Here, structures of type I-FHNH and I-EHNH Cascade complexes at different states are reported. •In type I-FHNH Cascade, Cas8fHNH loosely attaches to Cascade head and is adjacent to the 5′ end of the target single-stranded DNA (ssDNA). Formation of the full R-loop drives the Cascade head to move outward, allowing Cas8fHNH to detach and rotate ∼150° to accommodate target ssDNA for cleavage. •In type I-EHNH Cascade, Cas5eHNH domain is adjacent to the 5′ end of the target ssDNA. Full crRNA-target pairing drives the lift of the Cascade head, widening the substrate channel for target ssDNA entrance. •Altogether, these analyses into both complexes revealed that crRNA-guided positioning of target DNA and target DNA-induced HNH unlocking are two key factors for their site-specific cleavage of target DNA. #highlights:- •Structures of type I-FHNH and I-EHNH Cascade complexes at different states. •Cas8fHNH and Cas5eHNH are adjacent to the 5′ end of the target ssDNA. •R-loop formation allows rotation of Cas8fHNH to accommodate target DNA for cleavage. •Cas5eHNH and Cas11e of type I-EHNH Cascade form the channel for target DNA cleavage.

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Here is my latest blog about: Accelerate Genomic Sequencing with Intel OneAPI - https://lnkd.in/g9TChUFc Are you ready to accelerate genomic sequencing to new heights? Look no further than Skrots and Intel OneAPI. With our advanced technologies and cutting-edge solutions, we can help you revolutionize the way genomic sequencing is performed. Genomic sequencing is the process of determining the complete DNA sequence of an organism's genome. This process is crucial for understanding genetic variations... Do Like & Share :)