Extract-N-Amp™ Plant for ASPB

Description

The Extract-N-Amp™ Plant PCR Kits contain all the reagents necessary to rapidly extract genomic DNA from plant leaves and amplify targets of interest by PCR.

• Novel – single-step extraction of genomic DNA to PCR
• Fast – tissue to PCR in 15 minutes
• Convenient – no long enzymatic digestions

The Extract-N-Amp Plant PCR Kit offers a novel Extraction Solution that eliminates the need for freezing of plant tissues with liquid nitrogen, mechanical disruption, organic extraction, column DNA purification, or alcohol precipitation. The product includes a specially formulated hot start PCR ReadyMix for amplification directly from the extract. Extract-N-Amp products are Sigma Advanced Technology certified.

Procedure

Genomic DNA is extracted from 0.5 to 0.7 cm plant leaf disks that have been cut with a standard paper punch and simply incubated in Extraction Solution at 95 °C for 10 minutes. An equal volume of Dilution Solution is added to the extract to neutralize inhibitory substances prior to PCR. A portion of the DNA extract is then added directly to the optimized PCR mix supplied.

Application

Perfect for genotyping.

Beneficial Features

• Starting Material – 0.5 to 0.7 cm plant leaf disk
• Speed – Extraction of genomic DNA for PCR in 15 minutes
• Flexible – Extract and amplify target DNA sequences from a variety of plant species (Figure 1)
• Specific – Hot Start antibody for highly specific PCR amplification of genomic DNA
• Safe – No phenol/chloroform or other hazardous material

Pick the formulation that is right for you

The Extract-N-Amp Tissue PCR Kits include a specially formulated hot start PCR ReadyMix for amplification directly from the extract.

The PCR ReadyMix comes in two formulations:

• REDExtract-N-Amp™ PCR Ready Mix – contains an inert dye that acts as a tracking dye and allows for convenient loading of PCR reactions onto agarose gels for analysis
• Extract-N-Amp PCR Ready Mix – no dye contained

Order Information

Kit Contents

REDExtract-N-Amp Kit

• Extraction Solution
• Dilution Solution
• REDExtract-N-Amp PCR Ready Mix
• 2 ml Tubes for Extraction

Sample Data

PCR analysis of genomic DNA extracted from 5 different plant species using Sigma's Extract-N-Amp Plant Kit.

Figure 1. Genomic DNA was extracted from 0.5 cm leaf disks that were cut using a standard paper punch. DNA was extracted using the Extract-N-Amp Plant PCR Kit in less than 15 minutes. All samples were then amplified using the specially formulated Hot Start PCR mix. The products were generated from a 30-cycle duplex reaction containing primers specific to plant chloroplast (upper band) and primers specific to Cannabis sativa DNA (lower band). MW ladder is 100, 200, 400 and 800 bp. Data provided by Andy Hopwood, Forensic Science Service, Birmingham, England.

Sequence was resolved on an ABI 310 from a purified, 645 bp corn leaf PCR product.

Figure 2. The PCR product was purified with the GenElute™ PCR Clean-Up Kit (Product No. NA1020). The DNA extraction and PCR were performed using Sigma's Extract-N-Amp Plant PCR Kit. The sequence was obtained using the same primers as for the original PCR.

Stability of Extract-N-Amp Plant Extracts.

Figure 3. Eight disks were punched from a corn leaf, and DNA was extracted according to the procedure in the Technical Bulletin for the Extract-N-Amp Plant Kit. Two 4 µl aliquots from each were analyzed immediately by quantitative PCR with SYBR® Green detection on an ABI Prism 7700. DNA standards for quantitative PCR were purified DNA prepared from corn leaf tissue with the GenElute Plant Genomic DNA Kit (Product No. G2N70). Half of the leaf extracts were stored at 4 °C (recommended storage conditions) and the other half at 37 °C (accelerated storage). Quantitative PCR was repeated after 1 week, 3 weeks, 6 weeks and 6 months from extracts at 37 °C. Results for storage at 37 °C are shown. The average of 2 replicate PCR assays from each extract is plotted. Error bars represent one standard deviation.

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