Independent Creative Scheme——μCaler Hybrid System Brought You into 2.5 hr for Whole Process！！

Whole genome sequencing is not enriched and the sequencing depth is generally not high, which can be used to discover new genomic abnormalities in patient prognosis or treatment strategies, and is often applied in basic biomedical research. While sequencing after targeted enriching partial regions of genomes helps obtain higher sequencing depth at a lower cost, thereby achieving higher detection specificity and sensitivity. Therefore, it is more suitable for clinical diagnosis. Targeted sequencing generally achieves the enrichment by means of multiplex amplification PCR or hybrid capture. Among them, the comprehensive performance of hybrid capture method is much better than that of multiplex amplification method, especially in better sequencing complexity and uniformity; capture DNA is directly proportional to Input, CNV can be detected simultaneously through "read depth" method; it is less affected by DNA quality and has high tolerance to cfDNA and low-quality FFPE wax block samples. However, the process of hybrid capture method is time-consuming, complex and greatly affected by operators, which limits its application in time-efficient scenarios, such as MRD detection.

To overcome the above shortcomings, we developed a completely new hybrid capture method  based on independent intellectual property rights, i.e.,

μCaler Hybrid System  (hereinafter referred to as: μCaler Technology)

μCaler probes design based on the principle of "conjugation effect", which can bind more firm and have higher specificity (Fig 1.).

Fig 1.Schematic Diagram of Working Principle in μCaler.

Performance

Simplified process for easier operation

The whole process of Hybridization and capture withμCaler only takes 2.5 hr. Even with a typical library preparation process of 4.5 hr, all the targeted capture before sequencing can be completed in 7 hr, which is close to the method of multi-amplification (Fig 2.).

Fig 2. The workflow o fμCaler vs traditional.

Ultra-stable of Capture Performance

When using relatively small panels (<100 Kb) for traditional hybridization capture, the probes usually capture non target regions, resulting in a low target rate, and the capture performance are also vulnerable to the level of operators. As shown in Fig 3., when different operators test the traditional M71 Panel and μCaler M71 Panel for targeting the same area in different batches, the On-target rate of μCaler remain above 75%, but the On-target rate of traditional ranges from 10% to 70%, with sharp fluctuations (Fig 3. A).

The capture uniformity of μCaler, whether 0.2x or 0.5x mean coverage, are much higher than that of traditional (Fig 3. B), which can also be seen from the Fold 80 value (Fig 3. C).

Fig 3. Comparison ofμCaler Panel and Traditional Panel.

A:On-target; B:Target covered; C:Fold 80 base penalty.

Note: The pre-library was prepared from Human Male Genomic DNA (Promega-male, G1471) with the same input amount. The targeting regions ofμCaler M71 Panel is identical to the traditional M71 Panel with theμCaler Hybrid System and NadPrep Hybrid Capture (Traditional) for hybrid capture, respectively. Sequencing: Illumina Novaseq 6000，PE150.

When captured with other panels with different coverage regions and GC content, the capture performance ofμCaler are very stable and have high data utilization (Fig 4.)

 Fig 4. Comparison of differentμCaler Panels.

Panel information：M27 (3.2 Kb，GC range 25-55%); M44 (5.3 Kb，GC range 25-70%); M50 (5 Kb，GC range 37-90%); M580 (58 Kb, GC range 19-99%).

Conclusion

The μCaler Hybrid System is a brand-new creative scheme for hybridization capture that provides ultra stable performance with Kb-level panels, and the whole process takes only 2.5 hr.

Nanodigmbio launched the first product with this μCaler technology -μCaler MRD Solution! With customized μCaler Custom Panel 3-day ultra-fast production service, to contribute to MRD rapid detection.

Please look forward to the next chapter μCaler for MRD!

For research use only. Not for use in diagnostic procedures.