Microzone, a leader in molecular biology solutions, is excited to announce an upcoming application note detailing the efficiency and performance of the cell2seq – 16S rRNA Microbial Identification Kit. Part of Microzone’s renowned Free From range, this kit provides reliable solutions for molecular biology workflows, free from solvents, columns, and harsh chemicals, ensuring safer, more eco-friendly processes. This application note showcases exciting discoveries and techniques, highlighting the amazing results you can achieve with Microzone’s innovative products.

The cell2seq – 16S rRNA Microbial Identification Kit consists of three key steps for 16S rRNA sequencing: sample preparation, high-fidelity PCR, and post-PCR clean-up.

• In the first step, sample preparation is performed using microLYSIS Plus, a reagent designed to lyse microbial cells and extract high-quality DNA efficiently.
• The second step involves high-fidelity PCR amplification of 16S rRNA gene regions using Microzone's MegaMixCrystal High-Fidelity Q5 MasterMix, ensuring precise and reliable amplification.
• The final step is post-PCR clean-up, where the microCLEAN reagent removes residual PCR products, optimising amplicon recovery and maintaining the integrity of the final sequences for accurate microbial identification.

These three steps work seamlessly together, delivering an efficient and robust workflow for 16S rRNA sequencing.

The application note highlights the simplicity, reliability, and robustness of the 16S sequencing workflow, offering compelling insights into:

• Time Efficiency: The application note showcases the time-saving benefits of the workflow, with the total time for the three steps under 2 hours and hands-on time under 15 minutes. This streamlined process reduces the time spent compared to traditional DNA extraction and PCR clean-up methods, enabling researchers and clinical laboratories to accelerate their 16S sequencing without compromising quality or accuracy.
• Superior Amplicon Recovery: The PCR clean-up protocol, using the microCLEAN reagent, ensures minimal sample loss. This is illustrated with gel electrophoresis images of amplicons derived from Acinetobacter baumannii, Escherichia coli, and Pseudomonas aeruginosa, showing the successful removal of PCR residues and no amplicon loss. Amplicon recovery was also confirmed by measuring sample concentration using a Qubit instrument before and after clean-up with microCLEAN.
• High-Quality Metrics: Sanger sequencing results show remarkable quality metrics achieved with the kit's workflow, such as superior Quality Scores (QS) and Continuous Read Length (CRL). Amplification of the V3-V4 regions of the 16S gene was performed using MegaMixCrystal High-Fidelity Q5 MasterMix, ensuring exceptional sequence fidelity.
• Comprehensive Analysis: The note delves into sequence trimming and consensus generation using Unipro UGENE software. Results were validated against the NCBI BLAST 16S database, ensuring precise identification of bacterial species from both colonies and broth cultures.

This in-depth application note not only highlights the efficacy of the cell2seq – 16S rRNA Microbial Identification Kit but also provides a practical guide for scientists aiming to streamline their workflows while maintaining the highest data quality standards.

Click here to download the application note today — an essential read for microbiologists, molecular biologists, and anyone in the field of microbial identification and genomics!