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Terra PCR Direct Polymerase Mix
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Terra PCR Direct Polymerase Mix
Tech Note

Amplifying a gene of interest from raw and processed meat samples

Terra PCR Direct outperforms other enzymes with samples containing higher levels of PCR inhibitors

Introduction Results Conclusions Methods

Introduction  

Terra PCR Direct Polymerase Mix contains an optimized DNA polymerase that allows for direct amplification from tissue sources or crude extracts, without the need for DNA purification. It can amplify DNA targets up to 4 kb as well as difficult templates (including those with a GC content greater than 70%)—and it enables DNA amplification in the presence of PCR inhibitors, growth serum, and other source material components.

Results  

Comparison with other PCR enzymes

Terra PCR Direct Polymerase demonstrates an exceptional ability to provide consistently strong amplification of a target gene (cox1) directly from crude samples—specifically, from different types of raw meat (beef, chicken, and pork) and processed meat (chicken dumplings, beef meatloaf, and roast pork)—when compared to other PCR enzymes (Takara Ex Taq hot-start version [HS] and PrimeSTAR HS, GXL, and MAX).

Terra PCR Direct Polymerase Mix (Lane 1 in each gel) provided strong amplification for all six sample types, including the raw beef sample (Panel A). Although the PrimeSTAR series of PCR enzymes (Lanes 3–6) shows good overall performance with the crude extracts in Panels B–F, none of the enzymes other than Terra are able to effectively amplify the raw beef sample (Panel A).

Comparing Terra PCR Direct to other PCR enzymes for amplification from crude samples.

Comparing the ability of Terra PCR Direct Polymerase Mix to amplify a cox1 target gene of interest from different raw and processed meat samples relative to the performance of other PCR enzymes. Lane 1: Terra PCR Direct Polymerase Mix. Lane 2: Takara Ex Taq HS polymerase. Lane 3: PrimeSTAR HS DNA Polymerase. Lane 4: PrimeSTAR GXL DNA Polymerase, standard protocol. Lane 5: PrimeSTAR GXL DNA Polymerase, high-speed protocol. Lane 6: PrimeSTAR Max DNA Polymerase. Lane M: pHY Marker.

Conclusions  

This experiment demonstrates that most meat extracts can be amplified by Takara Ex Taq HS and PrimeSTAR HS, GXL, and MAX DNA Polymerases, but Terra PCR Direct Polymerase Mix shows a higher tolerance to PCR inhibitors than the other enzymes, regardless of the type of sample used. Terra PCR Direct polymerase generates a strong amplification band for the cox1 gene from the crude raw beef extract, a rich source of iron, zinc, and other metal ions that can interfere with PCR reactions. The fact that the other polymerases showed little or no amplification of the raw beef sample suggests that Terra is the best choice for analyzing meat extracts, as well as other samples that contain high levels of PCR inhibitors.

Methods  

Alkali and heat were used to prepare crude DNA extracts from raw meats (beef, chicken, and pork) and processed meats (chicken dumplings, beef meatloaf, and roast pork). The extracts were prepared by heating the meat samples for 10 minutes at 95°C in the presence of sodium hydroxide, followed by neutralization and centrifugation. Various PCR enzymes were used to amplify a cox1 gene fragment from a 1-μl aliquot of each crude extract in a 20-μl reaction volume. After amplification, 3 μl of each reaction was analyzed on a 1% Agarose L03 Gel to compare the effectiveness of each enzyme in amplifying the different crude extracts.

Related Products

Cat. # Product Size License Quantity Details
639270 Terra™ PCR Direct Polymerase Mix 200 Rxns USD $172.00

A hot-start polymerase mix that is developed for direct PCR amplification from tissue samples, crude extracts, and dirty templates. It is ideal for amplifying short DNA targets (up to 2 kb), regardless of GC content or template purity. The polymerase mix is supplied with separate tubes of buffer (with Mg2+ and dNTPs) and Proteinase K. The inclusion of Proteinase K enhances gel detection of the PCR products.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Documents Components Image Data Resources

Back

Terra PCR Direct was used to amplify the cyclin D2 gene (Ccnd2, 0.5 kb; Lane 1) and the transferrin receptor gene (TfrC 2 kb; Lane 2) from 1 μl of mouse blood treated with either EDTA or heparin

Terra PCR Direct was used to amplify the cyclin D2 gene (Ccnd2, 0.5 kb; Lane 1) and the transferrin receptor gene (TfrC 2 kb; Lane 2) from 1 μl of mouse blood treated with either EDTA or heparin
Panel A. Terra PCR Direct was used to amplify the cyclin D2 gene (Ccnd2, 0.5 kb; Lane 1) and the transferrin receptor gene (TfrC, 2 kb; Lane 2) from 1 μl of mouse blood treated with either EDTA or heparin. Panel B. Terra PCR Direct was used to amplify the mouse Ywhaz1 gene (1 kb) directly from either a 1 mm tail or 1.5 mm2 ear biopsy. A 4 μl aliquot of each sample was mixed with gel loading buffer that either lacked or contained proteinase K (Lanes 1 and 2, respectively). The PCR products treated with proteinase K ran as expected, whereas those without proteinase K treatment got stuck in the wells. Panel C. Terra PCR Direct was used to amplify the cytochrome c oxidase gene (cox1; 0.5 kb) directly from 0.5 mm (Lane 1) and 1.2 mm (Lane 2) tomato or spinach leaf cuttings (made using hole punches).

Back

639270: Terra PCR Direct Polymerase Mix

639270: Terra PCR Direct Polymerase Mix
639271 Terra™ PCR Direct Polymerase Mix 800 Rxns USD $573.00

A hot-start polymerase mix that is developed for direct PCR amplification from tissue samples, crude extracts, and dirty templates. It is ideal for amplifying short DNA targets (up to 2 kb), regardless of GC content or template purity. The polymerase mix is supplied with separate tubes of buffer (with Mg2+ and dNTPs) and Proteinase K. The inclusion of Proteinase K enhances gel detection of the PCR products.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Documents Components Image Data Resources

Back

Terra PCR Direct was used to amplify the cyclin D2 gene (Ccnd2, 0.5 kb; Lane 1) and the transferrin receptor gene (TfrC 2 kb; Lane 2) from 1 μl of mouse blood treated with either EDTA or heparin

Terra PCR Direct was used to amplify the cyclin D2 gene (Ccnd2, 0.5 kb; Lane 1) and the transferrin receptor gene (TfrC 2 kb; Lane 2) from 1 μl of mouse blood treated with either EDTA or heparin
Panel A. Terra PCR Direct was used to amplify the cyclin D2 gene (Ccnd2, 0.5 kb; Lane 1) and the transferrin receptor gene (TfrC, 2 kb; Lane 2) from 1 μl of mouse blood treated with either EDTA or heparin. Panel B. Terra PCR Direct was used to amplify the mouse Ywhaz1 gene (1 kb) directly from either a 1 mm tail or 1.5 mm2 ear biopsy. A 4 μl aliquot of each sample was mixed with gel loading buffer that either lacked or contained proteinase K (Lanes 1 and 2, respectively). The PCR products treated with proteinase K ran as expected, whereas those without proteinase K treatment got stuck in the wells. Panel C. Terra PCR Direct was used to amplify the cytochrome c oxidase gene (cox1; 0.5 kb) directly from 0.5 mm (Lane 1) and 1.2 mm (Lane 2) tomato or spinach leaf cuttings (made using hole punches).


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Citations Citations: Terra Direct

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