The High Sensitivity Small DNA Fragment Analysis Kit was designed for the automated analysis of small, low concentration DNA samples. Able to size DNA fragments and smears from 50 bp – 1,500 bp with unparalleled accuracy, the kit is ideal for the evaluation of small RNA NGS libraries. Other applications include genotyping, the quantitative analysis of PCR amplicons, and microsatellites.
|Components||500 Sample Kit||1,000 Sample Kit|
|Small Fragment Separation Gel||240 mL (DNF-230-0240)||500 mL (DNF-230-0500)|
|Intercalating Dye||30 µL (DNF-600-U030)||30 µL x 2 (DNF-600-U030)|
|5X 930 dsDNA Inlet Buffer||125 mL (DNF-355-0125)||300 mL (DNF-355-0300)|
|5X Capillary Conditioning Solution||50 mL (DNF-475-0050)||100 mL (DNF-475-0100)|
|HS Small Fragment Diluent Marker Solution||2.4 mL x 5 vials (DNF-371-0003)||2.4 mL x 10 vials (DNF-371-0003)|
|HS Small Fragment DNA Ladder||100 µL (DNF-372-U100)||100 µL x 2 (DNF-372-U100)|
|0.25X TE Rinse Buffer||125 mL (DNF-497-0125)||125 mL (DNF-497-0125)|
|BF-25 Blank Solution||8 mL (DNF-300-0008)||8 mL (DNF-300-0008)|
|Sample Volume Required||2 µL|
|Diluent Marker Volume Required||22 µL|
|DNA Sizing Range||50 bp – 1,500 bp|
Ultra-Short Array: 50 bp – 700 bp ≤ 5%; 700 bp – 1500 bp ≤ 10%
Short Array: 50 bp – 900 bp ≤ 5%; 900 bp – 1500 bp ≤ 10%
|DNA Sizing Accuracy||± 5% or better|
|DNA Sizing Precision||2% CV|
|DNA Fragment Concentration Range||5 pg/µL – 500 pg/µL input DNA|
|DNA Smear Concentration Range||200 pg/µL – 5000 pg/µL input DNA|
|DNA Quantification Accuracy||± 25 %|
|DNA Quantification Precision||15 % CV|
|Maximum DNA Concentration||500 pg/µL per fragment; 5000 pg/µL total|
|Total Electrophoresis Run Time||33 min (Ultra-Short Array, 22-47); 40 min (Short Array, 33-55)|
Sizing and quantification of small DNA fragments necessitates the use of a DNA ladder with precise, well selected fragments provided at a proper molarity. Figure 1 depicts a typical separation of the High Sensitivity Small Fragment DNA Ladder (DNF-372) using the High Sensitivity Small DNA Fragment Analysis Kit (DNF-477). Electrophoresis was performed on a Fragment Analyzer equipped with a Short Capillary Array (33-55), providing an excellent separation of the ladder fragments.
Small RNA NGS libraries are accurately assessed using the HS Small DNA Fragment Analysis Kit, promoting quality sequencing results. A small RNA NGS library is shown in Figure 2. Prior to electrophoresis the library was size selected, removing the appearance of a traditional library smear.
The High Sensitivity Small Fragment DNA Analysis Kit (DNF-477) is suited to any application that requires accurate and precise sizing and quantification of small DNA fragments and smears. Sheared DNA smears are sized and quantified with the same level of accuracy and precision as DNA fragments. The Smear Analysis function in PROSize® provides enhanced measurements over the default fragment analysis function. In Figure 3, the Smear Analysis region is bounded by the dotted red lines and the average size of the smear is listed in red numbers.
In addition to excellent sizing, the High Sensitivity Small Fragment Analysis Kit (DNF-477) also provides accurate and precise quantification. Figure 4 shows the results of measuring the same samples via two methods, a fluorometric instrument and the Fragment Analyzer. The measured concentrations were plotted against each other revealing a one-to-one linear relationship between the methods with a high coefficient of determination, indicating equivalent measurements by both instruments.