gel function using gene construction kit

3, upper panels), and these sIgM (−) populations were further isolated by fluorescence-activated cell sorting (FACS). Cμ guides 1, 2, and 3 generated 5.9, 11.7, and 9.2% sIgM (−) populations 2 weeks after transduction, as estimated by flow cytometry analysis ( Fig. These lentiviral clones were transduced into the AID −/− DT40 cell line, which constitutively expresses cell surface IgM (sIgM) because of the absence of Ig gene conversion ( 12). 2D) were further tested to functionally validate the guide sequences in the library. 100 ng of each purified DNA fragment were electrophoresed again at 100 V for 20 min (Figure 2).Three guide sequences specific to Cμ ( Fig. The fragments were purified with the FastGene ® Gel/PCR Extraction Kit. The target fragments were excised out of the gel and transferred in a 1.5 mL tube.

The 0.7% agarose gel was produced using 1x TAE buffer (Figure 1). The restriction digest was analysed by agarose gel electrophoresis at 100 V for 20 min. The customer also highlighted the fast preparation, easy handling, high recovery rate for large fragments and the unproblematic performance in downstream applications.Ī 6.9 kb large plasmid was digested with a restriction enzyme. In this AppNote the FastGene® Gel/PCR Extraction Kit was used for the isolation of two DNA bands after restriction digestion.īoth fragments show a good recovery rate after extraction. It is a well-known problem that the recovery of DNA fragments larger than 1 kb proves to be difficult and leads to the loss of large amounts of DNA. Lane 5,6 and 7: FastGene ® Gel/PCR Extraction Kit Lane 1: Unpurified PCR fragment M = Marker DNA was eluted in 20 µL elution buffer and 10 µL of eluate was analyzed on a 2% TAE agarose gel. After electrophoresis the DNA was purified using a competitor kit and the FastGene ® Gel/PCR Extraction Kit, according to manufacturers protocol. The DNA recovery from agarose gels using FastGene ® Gel/PCR Extraction Kits is very high even for small fragments.įigure: 10 µL of a PCR stock solution with a 100 bp amplicon were applied on a TBE agarose gel. 5 μL of eluted DNA were analyzed on a 1.5% TAE agarose gel. The figure demonstrates that the FastGene ® Gel/PCR Extraction Kit shows up to 90% of DNA recovery.įigure: PCR fragments of 300 bp were purified from 40 μL of a PCR stock solution using FastGene ® Gel/ PCR Extraction Kit and a competitor kit, according to manufacturers protocol. Everything you need to cut out your DNA fragments. Extra benefitsĮach FastGene ® Gel/PCR Extraction Kit includes 5 FastGene ® Agarose Gel Band Cutters and 50 µL Midori Green Advance DNA stain (enough to stain 1 L of agarose gels). With a simple and fast bind-wash-elute procedure you can purify DNA ranging from 15 bp to 10 kb with an elution volume of 20-50 µL. The FastGene ® Gel/PCR Extraction Kit provides spin columns, buffers, and collection tubes for silica-membrane-based purification of DNA fragments from agarose gels and PCR products. DNA fragments purified with FastGene ® Gel/PCR Extraction Kits are ready for direct use in all common downstream applications, like sequencing, ligation and transformation, restriction digestion, microarray analysis, PCR and in vitro transcription.

The FastGene ® Gel/PCR Extraction Kit is designed for the extraction of DNA from agarose gels and for the purification of PCR products. Details Two in one – DNA cleanup from agarose gels and PCR

Gel function using gene construction kit