Overview

Norgen’s Urine DNA Isolation Kit for Exfoliated Cells or Bacteria is designed for the rapid isolation of either: 1) human genomic DNA from exfoliated cells that have been shed into the urine from the urinary tract; or 2) bacterial genomic DNA from urine samples. The kit allows for the isolation of DNA from 1 to 50 mL of urine. The genomic DNA isolated from exfoliated cells can be used in a number of diagnostic and research applications including the diagnosis and monitoring of bladder, kidney, or other urinary-tract cancers. Bacterial genomic DNA from both human urine samples and urine samples from animals can be isolated with this kit in order to study the levels and types of bacteria that are present. The kit allows for the isolation of genomic DNA from both Gram negative and Gram positive bacteria, including E. coli, Proteus spp., Klebsiella spp., Enterobacter spp., Serratia spp., Pseudomonas spp, Clostridial ssp. and Leptospirosis spp.Chlamydia trachomatis and Neisseria gonorrhoeae.

Typical yields of human genomic DNA from exfoliated cells will vary depending on the cell density of the urine sample, which is affected by a number of factors including health, diet and sex of the individual donating the urine. Typical yields of bacterial genomic DNA will vary depending on the urine sample and the bacterial species, if any, present in the urine. Healthy humans generally have < 10, 000 CFU of bacteria per mL of urine, and this kit is sufficiently sensitive to isolate and detect DNA from even this minimal amount of bacteria. The genomic DNA purified with this kit is fully digestible with all restriction enzymes tested, and is completely compatible with downstream applications such as PCR, qPCR and Southern Blot analysis.

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Supporting Data

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Figure 1. Isolation and Detection of Biologically Active Genomic DNA from 1 mL of Urine. Genomic DNA was isolated from the exfoliated cells found in a 1 mL urine sample using Norgen's Urine DNA Isolation Kit for Exfoliated Cells or Bacteria. The purified DNA was eluted in 100 µL of Elution Buffer, and 5 µL of the isolated DNA was used as a template in quantitative PCR using human beta actin gene primers to detect the genomic DNA using the iQ SYBR Green Supermix (BioRad, #170-8882). The red lines in the PCR baseline graph above correspond to DNA standards, while the blue line corresponds to the successful PCR results when DNA isolated from the exfoliated cells in 1 mL of urine was used as the template.

Figure 2. Isolation and Detection of Bacterial Genomic DNA from a 1 mL Urine Sample. Bacterial genomic DNA was isolated from a 1 mL urine sample from a healthy individual using Norgen's Urine DNA Isolation Kit for Exfoliated Cells or Bacteria. The provided protocol was followed, and the purified urinary bacterial DNA was eluted in 100 µL of Elution Buffer. Five microliters of the eluted DNA was then used as a template in a quantitative PCR reaction to detect the bacteria using the iQ SYBR Green Supermix (BioRad, #170-8882). Healthy humans generally have <10, 000 CFU per mL of urine, and this kit is sensitive enough to isolate genomic DNA from this small amount of bacteria (blue line in graph above). The red lines in the above graph correspond to DNA standards.

Figure 3. Isolation of Genomic DNA from both Gram-Positive and Gram-Negative Urine Bacteria. Norgen's Urine DNA Isolation Kit for Exfoliated Cells or Bacteria was used to isolate genomic DNA from 1mL urine samples spiked with the Gram-positive bacteria B. cereus (1 million cells/mL) (Lanes 1 and 2), and the Gram-negative bacteria E. coli (1 million cells/mL) (Lanes 3 and 4). The provided protocol was followed, and the bacterial DNA was eluted in 100 µL of Elution Buffer. One tenth from each elution was loaded on 1.2% agarose gel. Lane M is Norgen’s MidRanger 1kb DNA Ladder. As it can be seen, genomic DNA could be successfully isolated from both Gram positive and Gram negative bacteria.

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