FAQ

Q1. If we buy one Seegene size marker, how can we use for 100 years?

A1. Unlike other commercial size markers which are consumable, we provide 12 clones (plasmids) corresponding to each base pair. Thus, the user can keep producing size markers by PCR using these clones.

Q2. What reaction quantity of template do you provide?

A2. We provide sufficient templates (12 plasmids) and primers, enough for 100 reactions. If needed, they can be transformed to E.coli in order to yield an unlimited supply of templates. Each PCR reaction generates 300~400 loadings.

Q3. What amount of primers do you provide?

A3. We also provide sufficient primers. Additionally, we provide the primer sequence in our product manual so that the user can synthesize the primer independently. Further, the user can purchase this primer from Seegene separately.

Q4. What are the main components for this kit?

A4. In case of "Forever 100bp Ladder Personalizer", we supply twelve plasmids corresponding to twelve sizes (100~1000bp, 1200bp, and 1500bp), one primer pair, and one loading dye, which are in fifteen tubes. In case of "Forever Multi-Ladder Personalizer", we provide the same amount of plasmids as the "Forever 100bp ladders", and two primers: one for 100bp increment and another for 50bp increment, and one loading dye, which are in seventeen tubes.

Q5. Does each base pair require a different PCR condition?

A5. No, each base pair requires the same PCR condition. Since this system is different from multiplex PCR, each base pair requires an individual PCR. To obtain a ladder of 12 sizes, the user has to amplify 12 tubes each having a different size of plasmid and then combine them together.

Q6. What is the price for this kit?

A6. In case of the USA and other countries where Seegene does not have an exclusive distributor, the end user price for "Forever 100bp Ladder Personalizer" is USD350 and the price for "Forever Multi-Ladder Personalizer" is USD400.

Q7. How can we make a 500bp or 1000bp band show a strong intensity?

A7. To obtain more intensive bands, you can increase the volume of the corresponding bands after PCR reactions by 3 times when mixing.

Q8. I want to transform the clones into E.coli. which antibiotics/ selection marker/ resistance marker?

A8. We use ampicillin. 100 micrograms of ampicillin are added to every 1 ml of LB broth. One microlitre of each plasmid should be used for transformation of E.Coli cells. The vector we use has high copy numbers. You could follow the instruction in the Molecular cloning, the method of "Heat shock". Among all steps, the most critical factor is "competent cell" to generate high efficiency.

Q9. I am getting ready to run the PCR reaction for the first time and am running into a problem. Our 10X PCR buffer contains 1.5 mM MgCl2 yet the protocol says to add 2.5 mM separately. Can I just supplement my 10X buffer by adding an additional 1mM to the reaction? What is the purpose of doing it separately?

A9. The final concentration of 2.5 mM MgCl2 is recommended, but this concentration could be adjustable, depending on the commercial brand of Taq DNA polymerase. Please refer to each instruction of Taq DNA polymerase.

Q10. How does it work?

A10. We supply the 12 different plasmids as ready-to use conditions for PCR amplification. It means that you can directly use the plasmids as templates for PCR amplification.

Q11. Is it easy to generate this ladder system after purchase?

A11. We already adjusted the concentration of each plasmid for direct use in PCR. The step you need is that you can amplify the 12 different fragments using the plasmids and mixing them.

Q12. Do we need to do 12 transformations and in each case isolate and confirm the positive clones?

A12. You do not need the steps of transformation, plasmid preps, restriction analysis, and so on.

Q13. How much DNA will I receive if I order this ladder system?

A13. Sufficient template and primers are supplied to provide enough markers for about a 20 years supply in the typical laboratory. If needed, the plasmids can be transformed to yield an unlimited supply of templates.

Q14. How can I make my own ladder?

A14. To create your own ladder, simply use the two included primers and 12 individual templates to amplify each of 12 differently sized fragments, quantitate, and mix according to your requirements.