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By combining the stringency of CTAB lysis with the speed and ease-of-use of silica-membrane purification, ISOLATE II Plant DNA Kit provides a fast method for the purification of high-quality genomic DNA from most plant cells, particularly those rich in polysaccharides, including leaves, bark, roots and fruits as well as dung, animal-fecal, soil and compost samples.
Some plant tissues do not lyse well in CTAB and so an SDS-based lysis buffer is also provided as an alternative. Residual amounts of RNA remaining can be removed using RNase treatment during lysis with the RNase A supplied in the kit.
ISOLATE II Plant DNA Kit shows excellent recovery of plant DNA when different homogenization techniques are used. The silica membrane in the columns is optimized to improve DNA binding to give high-yields of high-quality DNA, even from difficult samples such as freeze-dried budding leaves. The extracted DNA is ready to use for subsequent downstream reactions such as PCR.
ISOLATE II Plant DNA Kit has been designed to deliver optimal performance in qPCR in tandem with SensiFAST Real-Time PCR Kits and in end-point PCR with MyTaq DNA Polymerase.
|
Reagent |
10 Preps |
50 Preps |
250 Preps |
|
ISOLATE II Filters |
10 |
50 |
250 |
|
ISOLATE II Plant Columns |
10 |
50 |
250 |
|
Collection Tubes (2 mL) |
20 |
100 |
500 |
|
Lysis Buffer PA1 |
5 mL |
25 mL |
125 mL |
|
Lysis Buffer PA2 |
4 mL |
20 mL |
100 mL |
|
Precipitation Buffer PL3 |
1 mL |
10 mL |
25 mL |
|
Binding Buffer PB |
6 mL |
30 mL |
125 mL |
|
Wash Buffer PAW1 |
6 mL |
30 mL |
125 mL |
|
Wash Buffer PAW2 |
6 mL |
25 mL |
50 mL |
|
Elution Buffer PG |
13 mL |
13 mL |
30 mL |
|
R |
Both lysis buffers PA1 and PA2 can possibly become cloudy, if they are stored under 20°C. They contain CTAB (PA1) and SDS (PA2) which can precipitate. After a short incubation at 37°C for several minutes they should become clear again.
For the optimal recovery of large RNA/DNA fragments it is necessary to optimize the elution step. To increase the recovery rate it would be helpful to use a larger volume for elution, incubate the column with the elution buffer prior to centrifugation and use repeated elution steps. To avoid excessive dilution it may be helpful to reapply the eluted fraction again. Furthermore it could be helpful for DNA elution to heat the elution buffer to 70°C.
