Genetic Engineering & Biotechnology News

MAY1 2015

Genetic Engineering & Biotechnology News (GEN) is the world's most widely read biotech publication. It provides the R&D community with critical information on the tools, technologies, and trends that drive the biotech industry.

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Genetic Engineering & Biotechnology News | GENengnews.com | MAY 1, 2015 | 23 OMICS with micro-ball ended fbers for emission and excitation. These provide for higher detec- tion sensitivities (0.1 ng/µl) and signifcantly reduced background noise (for improved signal to noise ratio). This system uses a low cost, powerful light-emitting diode (LED) in- duced fuorescent detection system to excite at 520 nm and detects 590–650 nm emission wavelengths. Simple System Operation Buffer(s), markers, and samples are placed in the buffer/sample tray within the instrument and the capillary gel cartridge is inserted (Figure 3). Within the software, the user selects their preprogrammed method or creates a new program. The user enters run parameters and sample locations within the 96-well sample tray and begins the analysis. The Q-Analyzer software automatically cal- culates the size of detected fragments (bp) us- ing a reference DNA Ladder. Proofs Qsep100 was evaluated for use in geno- typing fragment analysis and for its limit of detection and linear performance and resolu- tion (Figure 4). Figure 4A shows the limit of detection (LOD) using a 576 bp PCR DNA fragment diluted in water vs. a sizing ladder. Sample concentrations range from 0.002 ng/µL to 0.01 ng/µL. Figure 4B shows linearity of detection with the same 576 bp PCR fragment serially diluted in a sample buffer. Separation condi- tions for Figures 4A and 4B are an injection voltage of 4 kV/10 s, a separation voltage 6 kV at ambient temperature. Figure 4C demonstrates the capability of the PCR and RFLP methodologies in de- termining co-infections of different human papilloma virus (HPV) types. The sample was determined to have a high risk genotype HPV (type 66) and a low risk genotype HPV (type 6). The conditions are an injection at 4 KV/4sec and a separation at 8 KV/220 sec- onds at ambient temperature. Figure 4D shows the resolution power of the system. Summary The Qsep100 CGE system is a robust, easy-to-use, cost-effective, and effcient al- ternative to traditional CGE and slab gel electrophoresis for DNA fragment analysis. Its unique multi-use, pen-shaped, capillary gel cartridge coupled with real-time LED- induced fuorescence detection eliminates the need for expensive cooling systems, main- tains high separation performance, and re- duces instrument and sample analysis costs. PCR and RFLP genotyping can be per- formed in an automated fashion at higher speeds (90% time saving) and lower costs ($0.11 to $0.20 per sample). With demonstrated resolution as low as 2–4 base pairs, a limit of detection as low as 0.1 ng/µl, and a linear performance of R2= 0.9997, it has been shown that Qsep100 is a powerful and cost-effective tool for genotyping applications including restriction digests, PCR product analysis, and total RNA and plasmid purifcation for both low- and high-through- put DNA and RNA research facilities. Varoujan D. Amirkhanian (varoujamir khanian@bioptic.com.tw ) is director and Shou-Kuan (Eric) Tsai (eric.tsai@bioptic. com.tw) is president at BiOptic, Zsolt Ronai M.D., Ph.D., is professor and research sci- entist at Semmelweis University, and Gene W. Stewart (gstewart@biophoretics.com) is president at BiopHoretics.Websites: www. biophoretics.com and www.bioptic.com.tw. Note: Qsep100 DNA Fragment Analyzer is distributed by BiopHoretics. Tutorial Figure 4. (A) Limit of detection. (B) Linearity of performance. (C) Genotyping results showing the capability of the PCR/RFLP methodology in determining co-infections of diferent HPV types. (D) Resolution. A B C D

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