Summary of the February 2013 issue of BioTechniques
In February 2013, BioTechniques will feature articles highlighting several new methods, including: (i) a new polymerase chain displacement reaction that enhancing sensitivity of qPCR, (ii) a novel approach in the analysis of large genomic deletions using next-generation sequencing, (iii) a software platform that enables the quantitative analysis of collective cell migration patterns, (iv) a high-throughput approach to screen for gain-of-function human microRNAs, and (v) a specially designed phase changing peptide that allows for more efficient bioseparations.In addition to the research article content in February, the issue will also contain a special Tech News feature exploring next-generation sequencing’s path to the $1000 genome as well as Citations highlighting recent trends in methods literature, the Troubleshooting Forum, and our popular Scientist Profile.
When it comes to exploring genomes, copy number variations, SNPs and insertions/deletions are critical landmarks. While sequencing today is faster and deeper than ever before, defining a large-scale genomic deletion, along with exact breakpoints, is not an easy task for users of NGS systems. The reason for this is that NGS systems, while producing millions of reads, generally produce short read lengths. However, as a team from Switzerland report in the February issue of BioTechniques, both the single molecule system of Pacific Biosciences as well as the Illumina NGS platform can be used for deletion mapping. The authors demonstrate the ability to map deletions from several kilobases up to several megabases, and define a set of parameters that can be used to align and determine break points for these deletions. While care must still be taken when dealing with cases where identical sequences flank the breakpoint, the approach presented here should enable more efficient and faster deletion analysis in the future.
Cell migration is crucial for many biological and developmental processes. Analyzing patterns of cell migration, especially collective migration patterns, can be useful in determining migration cues as well as the molecular mechanisms behind cell migration. To this end, a team of researchers report a new software platform for the analysis of cell migration patterns. This tool is capable of analyzing the width of streams of cells rapidly, providing researchers the chance to study correlated cell movements and assess the molecular signals involved in such migrations.
Quantitative PCR (qPCR) is a sensitive approach for the determination of sample copy number. Amplification relies on 2 primers surrounding a sequence of interest. But what if instead of two primers, four primers were used? And what if the polymerase had stand displacement activity? This is the idea behind the polymerase chain displacement reaction (PCDR) developed by a team of researchers and described in the February issue of BioTechniques. In PCDR, four primers are employed in the reaction – amplification is initiated from both the outer primers and the inner primers and by using a polymerase with strand displacement activity, PCDR enables increased template amplification per cycle compared to the standard two primer reaction and thus enhanced sensitivity in qPCR applications.
Identifying microRNAs that play a role in human disease can be a challenge. Although transcriptome analysis can identify differentially expressed microRNAs, their specific roles in disease progression and biology are unclear. In an attempt to design an assay that can identify differentially expressed microRNAs and their effects, a team of researchers from the Johns Hopkins University created a methodology incorporating the lentiviral expression of different microRNAs with qPCR to assess functionally related increases in specific microRNA abundance. The approach, detailed in an article slated for the February issue of BioTechniques, provides researchers with a new high-throughput methodology for assessing microRNAs and their cellular roles.
The $1000 genome continues to be a goal for many in the sequencing community. But how realistic is this? What is the cost associate with data storage and analysis? How much is the researcher’s time worth? In a special Tech News in February, contributing writer Jeffrey Perkel explores the landscape of genome sequencing, and poses the question: we will ever hit the $1000 genome, and what could hold us back.
Keywords: genomics, qPCR, miRNA, lentiviral vectors, next-generation sequencing, bioinformatics, genome analysis, CNV, SNP analysis, polymerase, cell migration/invasion, peptides, bioseperations, protein analysis