NanoHDA： A nanoparticle-assisted isothermal amplification technique for genotyping assays

Isothermal methods, such as helicase-dependent amplification （HDA）, have an advantage over polymerase chain reaction for DNA amplification owing to their ease of operation. Here, we developed a new HDA method that is nanoparticle-assisted, termed nanoHDA. This method uses gold nanoparticles （AuNPs） to improve the sensitivity and specificity of the isothermal method. In HDA, the denaturation of DNA templates is mediated by helicases, but this method is limited by the low denaturation efficiency of helicases. In this report, AuNPs with preferential affinity for single-stranded DNA （ssDNA） were utilized to improve the denaturation efficiency of helicases. The same affinity property of nanoparticles can also enhance specificity by suppressing primer-dimer formation. This nanoHDA method was employed to genotype the KRAS gene in genomic DNA samples from colorectal cancer patients, as achieved by the hybridization of nanoHDA amplicons using the NanoBioArray chip.

A Rapid and Cost-Effective Method for Genotyping Genome-Edited Animals：A Heteroduplex Mobility Assay Using Microfluidic Capillary Electrophoresis

The recent emergence and application of engineered endonucleases have led to the development of genome editing tools capable of rapidly implementing various targeted genome editions in a wide range of species.Moreover,these novel tools have become easier to use and have resulted in a great increase of applications.Whilst gene knockout（KO） or knockin（KI） animal models are relatively easy to achieve,there is a bottleneck in the detection and analysis of these mutations.Although several methods exist to detect these targeted mutations,we developed a heteroduplex mobility assay on an automated microfluidic capillary electrophoresis system named HMA-CE in order to accelerate the genotyping process.The HMA-CE method uses a simple PCR amplification of genomic DNA（gDNA） followed by an automated capillary electrophoresis step which reveals a heteroduplexes（HD） signature for each mutation.This allows efficient discrimination of wild-type and genome-edited animals down to the single base pair level.

Diagnostic Performance of the GenoType MTBDRplus and MTBDRs/Assays to Identify Tuberculosis Drug Resistance in Eastern China

Background： The WHO recently has recommended the GenoType MTBDRph/s version 1.0 and MTBDRs/version 1.0 assays for widespread use in countries endemic with drug-resistant tuberculosis. Despite this, these assays have rarely been evaluated in China, where the burden of drug-resistant tuberculosis is among the highest globally. Methods： Mycobacterium tuberculosis clinical isolates were obtained between January 2008 and December 2008. Isolates were tested for drug resistance against rifampicin （RFP） and isoniazid （INH） using the GenoType MTBDRplus assay and drug resistance against ethambutol （EMB）, ofloxacin （OFX）, and kanamycin （KM） using the Genotype MTBDILsl assay. These results were compared with conventional drug-susceptibility testing （DST）. Results： Readable results were obtained from 235 strains by GenoType MTBDRphts assay. Compared to DST, the sensitivity of GenoType MTBDRplus assay to detect RFP, INH, and multidrug resistance was 97.7%, 69.9%, and 69.8%, respectively, whereas the specificity for detecting RFP, INH, and multidrug resistance was 66.7%, 69.2%, and 76.8%, respectively. The sensitivity and specificity of the GenoType MTBDRsl assay were 90.9% and 95.2% for OFX, 77.8% and 99.5% for KM, 63.7% and 86.4% for EMB, respectively. Mutations in codon S531L of the rpoB gene and codon S315T1 ofKatG gene were dominated in multidrug-resistant tuberculosis （MDR-TB） strains. Conclusions： In combination with DST, application of the GenoType MTBDRplus and MTBDRsl assays may be a useful supplementary tool to allow a rapid and sale diagnosis of multidrug resistance and extensively drug-resistant tuberculosis.