Bacterial Genomic DNA Isolation Kit
For the rapid preparation of genomic DNA from bacteria
For research use only and NOT intended for in vitro diagnostics.
Bacterial Genomic DNA Isolation Kit
For the rapid preparation of genomic DNA from bacteria
Register today to receive an exclusive 15% off* on your first order.
Features and Benefits
- Isolate genomic DNA from all types of bacteria (both Gram-positive and Gram-negative)
- Rapid and convenient spin column protocol
- 96-well format available for high throughput
- High yield, high quality DNA for sensitive downstream applications including sequencing, PCR, qPCR and more
This kit is designed for the rapid spin column preparation of genomic DNA from 2 x 109 viable bacterial cells (between 0.5 and 1.0 mL of culture). This kit can be used for both Gram-negative and Gram-positive bacteria including Escherichia coli and Bacillus cereus. Purified genomic DNA is of an excellent quality and yield, and is fully compatible with restriction enzyme digestions, sequencing, PCR, qPCR and more. Also available in 96-well format for high throughput applications.
Details
Supporting Data
Kit Specifications (Spin Columns)
|
|
Input
|
2 x 109 bacterial cells
|
Column Binding Capacity
|
25 µg
|
Average Yield*
|
Up to 20 µg
|
Time to Complete 10 Purifications |
1 hour
|
* Yield will vary depending on the type of sample processed
Storage Conditions and Product Stability
All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years after the date of shipment. The kit contains a ready-to-use Proteinase K, which is dissolved in a specially prepared storage buffer. The buffered Proteinase K is stable for up to 2 years after the date of shipment when stored at room temperature.
Component | Cat. 17900 (50 preps) | Cat. 17950 (192 preps) |
---|---|---|
Resuspension Solution A | 20 mL | 60 mL |
Lysis Buffer P | 18 mL | 60 mL |
Solution BX | 28 mL | 110 mL |
Wash Solution A | 18 mL | 2 x 38 mL |
Elution Buffer B | 30 mL | 2 x 30 mL |
Proteinase K | 12 mg | 50 mg |
Spin Columns | 50 | - |
96-Well Plate | - | 2 |
Adhesive Tape | - | 4 |
Collection Plate | - | 2 |
Collection Tubes | 50 | - |
Elution Tubes (1.7 mL) | 50 | - |
Elution Plate | - | 2 |
Product Insert | 1 | 1 |
Documentation
FAQs
Spin Column, High Throughput
Column clogging can result from the following:
- The sample is too large.
Too many cells were applied to the column. Ensure that the amount of cells used is less than 2 x 109 viable cells, and that no more than 1 mL of culture is applied to the column/well. Clogging can be alleviated by increasing the g force and/or centrifuging for a longer period of time until the lysate passes through the column.
- Insufficient Vacuum (for High-throughput only).
Ensure that a vacuum pressure of at least -650 mbar or -25 in. Hg is developed.
- Centrifuge temperature is too low.
Ensure that the centrifuge remains at room temperature throughout the procedure. Temperatures below 15℃ may cause precipitates to form that can cause the wells to clog.
Experiencing gelatinous lysate prior to loading onto the column may be due to:
- The lysate/binding solution mixture is not homogeneous.
To ensure a homogeneous solution, vortex for 10-15 seconds before applying the lysate to the spin column.
- The sample is too large.
Too many cells are in the lysate preparation. Ensure that the amount of cells used is less than 2 x 109 viable cells, and that no more than 1 mL of culture is applied to the column/well.
A low genomic DNA yield may be caused by the following:
- The sample is old/overgrown.
The culture may have been overgrown, allowing lysis of older cells to occur more readily. This will lead to premature degradation of the genomic DNA. It may be necessary to use bacterial cultures before they reach maximum density.
- Incomplete lysis of cells.
Extend the incubation time of Proteinase K digestion or reduce the amount of bacterial cells used for lysis. Increase the lysozyme incubation time for gram-positive strains.
- The DNA elution is incomplete.
Ensure that centrifugation at 20,000 x g is performed after the 3,000 x g centrifugation cycle to ensure that all the DNA is eluted.
Sheared genomic DNA could be caused by:
- Improper handling of genomic DNA.
Pipetting steps should be handled as gently as possible. Reduce vortexing times during mixing steps (no more than 10-15 seconds).
- The cells are old.
Older cultures contain prematurely lysed cells which release endonucleases and can degrade DNA. Fresh cultures are recommended.
If the DNA does not perform well in downstream applications, it may be due to one or more of the following:
- DNA was not washed two times with the provided Wash Solution A.
Ensure the column/plate was washed two times with Wash Solution A. An additional wash with Wash Solution A can improve DNA performance in downstream applications, although it may reduce DNA yield.
- Ethanol carryover.
Ensure that the column/plate drying step after the Wash procedure is performed in order to remove traces of ethanol prior to elution. Ethanol is known to interfere with many downstream applications.
Yes, the DNA from this kit is good for both Nanopore sequencing and Long-read sequencing. Please contact our Tech support team at support@norgenbiotek.com and ask for reference publications.
Citations
Title | Isolation of a significant fraction of non-phototroph diversity from a desert Biological Soil Crust |
Citation | Frontiers in Microbiology 2015. |
Authors | U Nunes da Rocha, H Cadillo-Quiroz, U Karaoz, L Rajeev, N Klitgord, S Dunn, V Truong, M Buenrostro, BP Bowen, F Garcia-Pichel, A Mukhopadhyay, TR Norther, EL Brodie |
Title | Genotypic and Phenotypic Heterogeneity in Alicyclobacillus acidoterrestris: A Contribution to Species Characterization |
Citation | PLoS One 2015. |
Authors | Bevilacqua, A., Mischitelli, M., Pietropaolo, V., Ciuffreda, E., Sinigaglia, M., & Corbo, M. R Milena Sinigaglia1, Maria Rosaria Corbo1 |
Title | Characterization and the influence of milk solids-not-fat on the bacteriocin produced by Lactococcus lactis subsp.lactis S20 isolated from Chinese sauerkraut, a traditional fermented vegetable |
Citation | Annals of Microbiology 2015. |
Authors | Yih-Zhet Chin, Selvi Velu, Fatimah Abu Bakar, Mahmud-Ab-Rashid Nor-Khaizura |
Title | Genotypic and phenotypic virulence characteristics and antimicrobial resistance of Yersinia spp. Isolated from meat and milk products |
Citation | Journal of Food Science 2015. |
Authors | F Ozdemir, S Arslan |
Title | Molecular Epidemiology of Escherchia Coli, in HIV-Positive Individuals in South west Nigeria |
Citation | International Archives of Medicine 2015. |
Authors | LE Okoror, DC Fashina, TS Jimoh, EA Oisagah |
Title | Insertion/Deletion-Based Approach for the Detection of Eschercia coli O157:H7 in Freshwater Environments |
Citation | Environmental Science & Technology 2014. |
Authors | SY Wong, A Paschos, RS Gupta, HE Schellhorn |
Title | Sequence Analysis of CRISPR Arrays of Erwinia amylovora isolates from Canada |
Citation | Acta Horticulturae 2014. |
Authors | AI Yagubi, AJ Castle, AM Svircev |
Title | Functional validation of putative toxin-antitoxin genes from the Gram-positive pathogen Streptococcus pneumoniae: phd-doc is the fourth bona-fide operon |
Citation | Frontiers in Microbiology 2014. |
Authors | WT Chan, CC Yeo, E Sadowy, M Espinosa |
Title | Isolation and Molecular Characterization of Brucella Isolates in Cattle Milk in Uganda |
Citation | BioMed Research International 2014. |
Authors | DR Mugizi, S Muradrasoli, S Boqvist, J Erume, GW Nasinyama, C Waiswa, G Mboowa, M Klint, U Magnusson |
Title | Potential Immunogenic Polypeptides of Burkholderia pseudomallei Identified by Shotgun Expression Library and Evaluation of Their Efficacy for Serodiagnosis of Meliodiosis |
Citation | Journal of Medical Sciences 2013. |
Authors | Suat Moi Puah, SD Puthucheary, Kek Heng Chua |