What’s DNA size selection?
DNA size selection (or DNA sizing) is the targeted capture of DNA fragments of a specific size or a size range. It’s broadly used in sequencing. And it’s a sample preparation step that has the greatest impact on the quality of sequencing results. Ineffective sizing can waste sequencing capacity on low molecular weight material such as adapter-dimers or primer-dimers (see DNA Clean-up) and imprecise sizing can hurt data quality. Precise DNA sizing can boost sequencing efficiency, save money, improve data, and even allow sequencing of low-input samples.
Why is DNA size selection important?
There are two types of sequencers: short-read sequencers (Illumina and Ion Torrent) and long-read sequencers (Pacific Biosciences and Oxford Nanopore Technologies). Both benefit from precise sizing, in a slightly different way.
Short-read sequencers operate best when fed DNA libraries that contain fragments of similar sizes, often in a very specific range recommended by the manufacturer. When libraries are not properly size-selected, these sequencers become significantly less efficient. It may take two lanes of sequencing, to accomplish what could be done in a single lane with a well-sized library.
Long-read sequencers tend to produce very-long reads by fully sequencing a long DNA fragment. When fragments are short, the read is limited. So it’s desired to remove smaller fragments using size selection, which allows the sequencers to focus on the DNA fragments most amenable to producing the longest reads.
How does DNA size selection work?
Figure 1. DNA size selection workflow.
- Mix: Mix MagVigen™ nanoparticles with DNA/RNA sample.
- Bind: When mixed with a sample, MagVigen™ nanoparticles bind to the desired DNA/RNA size fragments. This interaction relies on multiple factors between the DNA/RNA molecules and the surfaces of the nanoparticles, such as charge-charge interactions, hydrophilic/hydrophobic interactions, Van der Waal and other intermolecular forces. NVIGEN engineers the surface chemistry of the Magnetic beads and the buffer environment to provide the most ideal interaction for desired DNA size fragment selection.
- Wash: The beads respond to a magnetic force (by use of a magnet), allowing bound material to be rapidly and efficiently separated from the rest of the sample. After unbound materials are removed by aspiration, the nanoparticle-bound targets are washed by using the magnet.
- Elute: The bound target is released from the nanoparticle and can be used for downstream applications.
SPRI beads-based protocols among the best methods
SPRI (solid-phase reversible immobilization) beads-based protocols don’t need column or centrifugation. They use magnetic force to aggregate, wash and elute targets. This makes the workflow simple and easy for automation, which leads to higher throughput.(Please see DNA Purification & Isolation Using SPRI and SPRI for the Isolation of PCR Products.)
While some tutorial source (e.g. BiteSizeBio‘s dna-sizing-tutorial) says beads-based protocols should not be used when more precise sizing is needed and they’re most useful in experiments where DNA is abundant, this knowledge is outdated.
The capability and capacity of any DNA size selection methods come from the functional groups assembled to beads, not directly from the beads themselves. Nanobeads, like MagVigen™, manufactured with the state-of-the-art nanotechnologies can be engineered at the molecular level to assemble a variety of functional groups. And nanobeads are in fractional (1/10) size of conventional beads, which creates 10x bigger surface area off the same amount of beads, and new advanced beads dynamics. These new properties have increased the capability and capacity of beads-based protocols to be among the best methods now.
Figure 2. 1µg Fisher DNA ladder was size-specifically captured by MagVigen™ nanobeads by adjusting the ratios of MagVigen™ volume vs. DNA sample volume. The numbers on top indicate the ratio.
Long bp size selection
One of the MagVigen™ nanobeads’ unique advantages is long bp size selection, such as 700bp, 1000bp, 1500bp, 3000bp, and 6000bp.
Figure 3. 1µg Fisher Mix Ladder was incubated with MagVigen™ DNA Size Selection kit (K61001-1500) following standard protocol. The resulting DNA was analyzed on a 3% agarose gel. DNA<1000bp was greatly reduced and long sizes of DNA>1000bp was largely retained at an average yield of 80%.
Figure 4. DNA sizing results of 700bp by MagVigen™ DNA Size Selection kit (K61001-700).
Improve your DNA size selection
We cordially invite you to try our state-of-the-art MagVigen™ DNA Size Selection Kits (K61001) for your DNA purification and size selection needs.
Or interested in sequence specific DNA/RNA capture?
Check our our MagVigen™-Streptavidin products (K61002).