DESCRIPTION

    TOROGreen^® Yeast colony PCR Kit is used to amplify fragments from the yeast genome as well as transformed plasmids. The same as E coli.colony PCR, PCR amplification directly from a yeast colony without DNA or plasmid extraction from yeast for the robust direct PCR mix. The protocol of this kit is easy, rapid and low price than the traditional method. TOROGreen^® 6×DNA loading buffer contains the fluorescence dye, which is directly detected on 470nm LED transillum inator after the DNA electrophoresis.

FEATURES

- Easy-to-use: the same as E coli.colony PCR without DNA or plasmid extraction.

- Economical: the protocol is easy, rapid and low price than the traditional method.

- High performance: can amplify 40kb genomic DNA and ＞90%  GC-rich target with 80-fold higher fidelity than Taq DNA polymerase.

APPLICATION

 - Yeast Colony PCR

COMPONENT

    The kit includes the following reagents, which can be used for 200 reactions for a total 10ul reaction volume.

   Cat No. ：DPK-100

   2× ASFastTM Direct PCR Mix                               1ml ×1tube

   TOROGreen^® 6×DNA loading buffer                1ml ×1tube

BRIEF PROCEDURE

PROTOCOL

1. Pick the yeast colony.

（1）Angle the 10ul sterile pipet tip down towards a colony and just barely touch the colony.

（2）Place the tip into 10 uL of sterile water in a 1.5ml clean microcentrifuge tube and pipet up and down several times. Label the tube with a colony number and store the yeast colony in water at 4℃.

2. Standard reaction setup.

（1）Prepare the 9ul Primer-PreMix on ice as the following components.

（2）Add 1ul yeast colony suspension into the 9ul Primer-preMix on ice, and pipet up and down several times. Gently mix the reaction solutions and spin down in microcentrifuge.      

3. Set up cycling conditions

4. Confirm the correct colonies by electrophoresis.

 (1) Add 1 volume of TOROGreen^® 6×DNA loading buffer to 5 volumes of PCR products.

 (2) Mix well, spin down and load 5ul of the mixture and 2ul of the suitable TOROGreen^® Loading Marker.

 (3) Run on agarose electrophoresis to detect PCR products and marker. No additional dye is required for the PCR products and marker.

 (4) Realtime observation of electrophoresis with a 470nm blue light source fitted to the electrophoresis tank and confirm the correct colonies.

5. Reconfirm the the correct colonies by sequencing .

 (1) Sequencing the PCR products to reconfirm the correct colonies.

 (2) Add the remaining 9ul yeast colony suspension into fresh YPD in a test tube, and grow it to saturation overnight.

STORAGE
     This reagent can be stored at 4°C for 1 months. For longer storage,  this reagent should be kept at -20°C

Ordering Information

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