The DNA purification procedure utilizes spin minicolumns with membranes which efficiently and selectively bind nucleic acids. In the first step the CB Buffer is added to a DNA sample. It causes proteins to degrade and enables DNA binding to the column membrane. As an added convenience, the binding buffer contains a colour indicator, which facilitates easy monitoring of the solution’s pH for optimal DNA binding. The two-step washing stage efficiently removes impurities and enzyme inhibitors. The purified DNA is eluted using either a low ionic strength buffer (Elution Buffer) or water (pH 7.0-9.0) and can be used directly in all downstream applications such as PCR, qPCR, Southern blotting, DNA sequencing, enzymatic restriction, ligation and so forth or stored until ready to use.
up to 100 μl of a DNA sample
90-99%, depending on DNA fragment length (in the range of 100 bp – 10 kb)
DNA FRAGMENT LENGTH
100 bp – 10 kb
DNA fragments in the 50-100 bp and 10-30 kbp range can also be purified, as can genomic and plasmid DNA, however the efficiency will be decreased.
approx. 25 μg DNA
A260/A280 ratio = 1.7-1.9
RECOMMENDATIONS AND IMPORTANT NOTES
The optimal volume of the elution buffer used should be chosen in line with the amount of DNA in the sample and to the final DNA concentration expected. The use of 30-100 μl of the Elution Buffer is recommended. If a high DNA concentration is desired, the elution volume may be reduced down to
20 μl. It should be noted that this may reduce the efficiency of the DNA retrieval. It is essential to apply the elution buffer precisely onto the centre of the membrane. In order to maximize the DNA retrieval heat the Elution Buffer to 80 ̊C and incubate it on the membrane for 10 minutes. If full DNA retrieval is required, a second elution should be performed. For second elution, repeat steps 10-13 of the Isolation Protocol (section XI), placing the purification column in a new, sterile 1.5 ml Eppendorf tube.
The Elution Buffer does not contain EDTA, which may interfere with some enzymatic reactions.
The CB Buffer contains an indicator, which enables pH monitoring. Yellow indicates that the pH of the solution is lower than 7.5, which guarantees optimal DNA binding to the membrane. When the pH is higher than 7.5, solution will turn pink. It may happen on the occasion, when the pH of a DNA sample considerably differs from the standard parameters of the DNA treatment operations (pH>9.0). In this case, it is essential to add 10 μl 3 M sodium acetate (pH 5.2). It will lower the pH, enabling the solution to bind efficiently to the minicolumn membrane.
EXTRACTME DNA CLEAN-UP KIT (EM07) - DISCONTINUED - Manual
EXTRACTME DNA CLEAN-UP KIT – TROUBLESHOOTING
The quality of each production batch (LOT) of the EXTRACTME DNA CLEAN-UP kit is tested using standard QC procedures. The purified DNA concentration, quality and stability are evaluated by gel electrophoresis and spectrophotometer.
EXTRACTME DNA CLEAN-UP KIT (EM07) - DISCONTINUED - CoA L. 746272
New nuclease from extremely psychrophilic microorganism Psychromonas ingrahamii 37: identification and characterization
Authors Maciejewska, N., Walkusz, R., Olszewski, M., & Szymańska, A Products EXTRACTME DNA BACTERIA KIT (EM02), EXTRACTME DNA CLEAN-UP KIT (EM07) – DISCONTINUED, EXTRACTME DNA GEL-OUT KIT (EM08) – DISCONTINUED, PWO Polymerases, Plasmid DNA Isolation Kits Year 2018 Source Molecular biotechnology, 61(2), 122-133.
Identification and characterization of single-stranded DNA-binding protein from the facultative psychrophilic bacteria Pseudoalteromonas haloplanktis
Authors Marcin Olszewski, Marta Nowak, Anna Cyranka-Czaja, Józef Kur Products EXTRACTME DNA BACTERIA KIT (EM02), EXTRACTME DNA CLEAN-UP KIT (EM07) – DISCONTINUED, EXTRACTME DNA GEL-OUT KIT (EM08) – DISCONTINUED, PWO Polymerases, PCR & Real-Time PCR Year 2014 Source Microbiological Research 169 (2014) 139– 147
Sequence, genome organization, annotation and proteomics of the thermophilic, 47.7-kb Geobacillus stearothermophilus bacteriophage TP-84 and its classification in the new Tp84virus genus.
Authors Piotr M. Skowron, Andrew M. Kropinski, Joanna Zebrowska, Lukasz Janus, Kasjan Szemiako, Edyta Czajkowska, Natalia Maciejewska, Malgorzata Skowron, Joanna Łoś, Marcin Łoś, Agnieszka Zylicz-Stachula Products EXTRACTME DNA CLEAN-UP KIT (EM07) – DISCONTINUED, PCR & Real-Time PCR Year 2014 Source PLOS ONE
Characterization of single-stranded DNA-binding proteins from the psychrophilic bacteria Desulfotalea psychrophila, Flavobacterium psychrophilum, Psychrobacter arcticus, Psychrobacter cryohalolentis, Psychromonas ingrahamii, Psychroflexus torquis, and Photobacterium profundum
Authors Marta Nowak, Marcin Olszewski, Marta Śpibida and Józef Kur Products EXTRACTME DNA BACTERIA KIT (EM02), EXTRACTME DNA CLEAN-UP KIT (EM07) – DISCONTINUED, EXTRACTME DNA GEL-OUT KIT (EM08) – DISCONTINUED, PCR & Real-Time PCR, PWO Polymerases Year 2014 Source Nowak et al. BMC Microbiology 2014, 14:91