Notice & News
The High Sensitivity Large Fragment Analysis Kit is used for the automated sizing and quantification of large DNA fragments and smears. Common applications include as a quality assurance step in long read sequencing, checking the quality of a DNA extraction, and evaluating the quality of degraded DNA samples. Ideal for low concentration samples, this kit is able to size samples through 20 Kb.
A quality ladder separation is important to the accurate quantitation and sizing of nucleic acid samples. The High Sensitivity Large Fragment DNA Ladder, as shown in Figure 1, provides such a separation when run on the Short Capillary Array (33-55) or the Long Capillary Array (55-80), allowing for the accurate quantification and sizing of large DNA samples. This ladder allows for quantification of fragments down to 0.005 ng/µL and sizing through 20,000 bp.
Figure 1. A typical separation of the High Sensitivity Large Fragment DNA Ladder (DNF-397) on a Fragment Analyzer equipped with a Short Capillary Array (33-55) using the High Sensitivity Large Fragment Analysis Kit (DNF-493).
Through the use of a high quality ladder, separation gel, and electrophoresis parameters, the High Sensitivity Large Fragment Analysis Kit (DNF-493) is able to accurately measure large DNA fragments and smears through 20 Kb. Large DNA fragments are sized with a high degree of accuracy. Figure 2 depicts the separation of a synthetic 15 Kb DNA fragment that was sized at 14.938 Kb, within 1% of the known base pair size.
Figure 2. Separation of a 15 Kb DNA fragment on a Fragment Analyzer equipped with a Short Capillary Array (33-55) using the High Sensitivity Large Fragment Analysis Kit (DNF-493)
Large DNA smear analysis allows for the evaluation of sheared genomic DNA (gDNA) and the assessment of DNA purification quality, among other things. Figure 3 depicts the separation of a 15 Kb DNA smear generated from the shearing of E. coli gDNA. An important feature of PROSize® is the Smear Analysis function which provides an accurate assessment of the smear size distribution. The Smear Analysis region, defined by the dotted red lines, displays the average size of the smear in red.
Figure 3. Separation of sheared E.coli gDNA performed on a Fragment Analyzer equipped with a Short Capillary Array (33-55) using the High Sensitivity Large Fragment Analysis Kit (DNF-493). DNA smears can be analyzed further in PROSize using the Smear Analysis function, indicated by the dotted red lines on the electropherogram. Smear Analysis functionsizing is shown in red numbers.
Precise and accurate measurements of large DNA smears and fragments will become increasingly important as molecular biology progresses and new applications develop. Next generation long read sequencing, synthesis of genetic constructs, and restriction digests are only some of the current applications that necessitate accurate measurements of large smears and fragments.