More than 99% of microorganisms on the earth are unculturable with known culturing techniques. The emergence of metagenomics with high-throughput sequencing technologies has enabled researchers to capture a comprehensive view of a complex bacterial community which comprises ...

With the advent of high-throughput sequencing technologies, multiple bacterial genomes can be sequenced in days. While the ultimate goal of de novo assembly of bacterial genomes is progressing, changes in the genomic sequence of closely related bacterial strains and isolates are now ea ...

Methods for in-depth characterization of transcriptomes and quantification of transcript levels have emerged as valuable tools for understanding cellular physiology and human disease biology, and have begun to be utilized in various clinical diagnostic applications. Today, ...

sRNA-Seq is an unbiased method that allows for the discovery of small noncoding RNAs in bacterial transcriptomes through direct cloning and massively parallel sequencing by synthesis. Small bacterial transcripts are enriched from a total RNA preparation and modified with 5′ and 3′ link ...

MicroRNAs are a class of small noncoding RNA molecules that play a pivotal role in the regulation of gene expression at the posttranscriptional level. Most large double-stranded DNA viruses, mainly the herpesvirus family, are known to express miRNAs. Viral miRNAs can regulate both viral- and ...

New high-throughput technologies continue to emerge for studying complex microbial communities. In particular, massively parallel pyrosequencing enables very high numbers of sequences, providing a more complete view of community structures and a more accurate inference of t ...

Next-generation sequencing (NGS) is a powerful tool that can be utilized to profile and compare microbial populations. By amplifying a target gene present in all bacteria and subsequently sequencing amplicons, the bacteria genera present in the populations can be identified and compa ...

The chaperonin-60 universal target (cpn60 UT) is generated from a set of PCR primers and provides a universally conserved, phylogenetically informative sequence signature for determining the composition of microbial communities by DNA sequencing. Pyrosequencing of cpn60 UT am ...

RNA-sequencing (RNA-Seq) is a digital display of a transcriptome using next-generation sequencing technologies and provides detailed, high-throughput view of the transcriptome. The first step in RNA-Seq is to isolate whole transcriptome from total RNA. Since large ribosomal RNA ( ...

Comprehensive evaluation of microbial diversity in almost any environment is now possible. Questions such as “Does the addition of fiber to the diet of humans change the gastrointestinal microbiota?” can now be answered easily and inexpensively. Tag-encoded FLX-amplicon pyrosequ ...

Metagenomics has evolved over the last 3 decades from the analysis of single genes and their apparent diversity in an ecosystem to the provision of complex genetic information relating to whole ecosystems. Metagenomics is a vast subject area in terms of methodology, which encompasses a suite ...

The digestive tract of animals contains a very large numbers of microorganisms with a high diversity. Traditionally, characterization of these microbial communities has relied on the ability to clonally culture each microorganism. With significant improvements in nucleotide ...

Metatranscriptomics has been developed to help understand how communities respond to changes in their environment. Metagenomic studies provided a snapshot of the genetic composition of the community at any given time. However, short-timescale studies investigating the respo ...

Whole-genome techniques toward identification of microbial genes required for their survival and growth during infection have been useful for studies of bacterial pathogenesis. The advent of massively parallel sequencing platforms has created the opportunity to markedly ac ...

Next-generation sequencing technologies have a massive throughput, which dramatically reduces the cost of sequencing per gigabase, compared to standard Sanger sequencing. To make the most efficient use of this throughput when sequencing small regions or genomes, we developed a ba ...

With rapid development of next-generation sequencing (NGS) technologies, it is becoming increasingly feasible to sequence entire genomes of various organisms from virus to human. However, in many occasions, it is still more practical to sequence and analyze only small regions of the ent ...

The library preparation step is of critical importance for the quality of next-generation sequencing data. The use of the polymerase chain reaction (PCR) as a part of the standard Illumina library preparation protocol causes an appreciable proportion of the obtained sequences to be dupl ...

DNA library preparation is a common entry point and bottleneck for next-generation sequencing. Current methods generally consist of distinct steps that often involve significant sample loss and hands-on time: DNA fragmentation, end-polishing, and adaptor-ligation. In vitro tr ...

Cytosine methylation is an epigenetic mark that has a significant impact on the regulation of transcription and replication of DNA. DNA methylation patterns are highly conserved across cell divisions and are therefore highly heritable. Furthermore, in multicellular organisms, D ...

The most polymorphic genetic markers are DNA regions composed of variable number tandem repeats (VNTRs) (1,2). Detection of the various VNTRs is possible by restriction fragment-length polymorphism (RFLP) analysis using the Southern blot procedure (3). However, this procedure is ti ...