Genome

About: Genome is a research topic. Over the lifetime, 74231 publications have been published within this topic receiving 3819713 citations.

High-throughput sequencing of DNA G-quadruplex structures in the human genome

Abstract: G-quadruplexes (G4s) are nucleic acid secondary structures that form within guanine-rich DNA or RNA sequences. G4 formation can affect chromatin architecture and gene regulation and has been associated with genomic instability, genetic diseases and cancer progression. Here we present a high-resolution sequencing-based method to detect G4s in the human genome. We identified 716,310 distinct G4 structures, 451,646 of which were not predicted by computational methods. These included previously uncharacterized noncanonical long loop and bulged structures. We observed a high G4 density in functional regions, such as 5' untranslated regions and splicing sites, as well as in genes previously not predicted to contain these structures (such as BRCA2). G4 formation was significantly associated with oncogenes, tumor suppressors and somatic copy number alterations related to cancer development. The G4s identified in this study may therefore represent promising targets for cancer intervention.

Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples

Abstract: Genome sequencing has become a powerful tool for studying emerging infectious diseases; however, genome sequencing directly from clinical samples (ie, without isolation and culture) remains challenging for viruses such as Zika, for which metagenomic sequencing methods may generate insufficient numbers of viral reads Here we present a protocol for generating coding-sequence-complete genomes, comprising an online primer design tool, a novel multiplex PCR enrichment protocol, optimized library preparation methods for the portable MinION sequencer (Oxford Nanopore Technologies) and the Illumina range of instruments, and a bioinformatics pipeline for generating consensus sequences The MinION protocol does not require an Internet connection for analysis, making it suitable for field applications with limited connectivity Our method relies on multiplex PCR for targeted enrichment of viral genomes from samples containing as few as 50 genome copies per reaction Viral consensus sequences can be achieved in 1-2 d by starting with clinical samples and following a simple laboratory workflow This method has been successfully used by several groups studying Zika virus evolution and is facilitating an understanding of the spread of the virus in the Americas The protocol can be used to sequence other viral genomes using the online Primal Scheme primer designer software It is suitable for sequencing either RNA or DNA viruses in the field during outbreaks or as an inexpensive, convenient method for use in the lab

Patterns of damage in genomic DNA sequences from a Neandertal

Abstract: High-throughput direct sequencing techniques have recently opened the possibility to sequence genomes from Pleistocene organisms. Here we analyze DNA sequences determined from a Neandertal, a mammoth, and a cave bear. We show that purines are overrepresented at positions adjacent to the breaks in the ancient DNA, suggesting that depurination has contributed to its degradation. We furthermore show that substitutions resulting from miscoding cytosine residues are vastly overrepresented in the DNA sequences and drastically clustered in the ends of the molecules, whereas other substitutions are rare. We present a model where the observed substitution patterns are used to estimate the rate of deamination of cytosine residues in single- and double-stranded portions of the DNA, the length of single-stranded ends, and the frequency of nicks. The results suggest that reliable genome sequences can be obtained from Pleistocene organisms.

Recombineering: a powerful new tool for mouse functional genomics

Abstract: Highly efficient phage-based Escherichia coli homologous recombination systems have recently been developed that enable genomic DNA in bacterial artificial chromosomes to be modified and subcloned, without the need for restriction enzymes or DNA ligases. This new form of chromosome engineering, termed recombinogenic engineering or recombineering, is efficient and greatly decreases the time it takes to create transgenic mouse models by traditional means. Recombineering also facilitates many kinds of genomic experiment that have otherwise been difficult to carry out, and should enhance functional genomic studies by providing better mouse models and a more refined genetic analysis of the mouse genome.