What are the best ways to reduce generation of chimeric molecules during PCR?

The chimeric molecules generated during PCR are really bad for our study on recombination, since the chimeras look exactly like recombination. I wonder what and how (would be better if protocol is...

06 July 2016 9,547 5 View

What is the average disance from a mismatched base for a base substitution occurring during mismatch repair?

Especially in mammalian cells.

05 June 2016 1,312 1 View

Would you like to share with me about isolation of single cells?

I am interested in all the methods or instruments used to isolate single cells especially from mammalian tissues. Any experience of you is appreciated.

09 October 2015 412 1 View

What percent of template can be synthesized during PCR?

For example, the parameters for PCR are denaturing at 95, annealing at 58 and elongation at 72, while the melting temperatures of primers are around 60.

04 May 2015 4,344 13 View

Does anyone have experience about the efficiency for linear DNA synthesis?

We are trying to synthesize mouse DNA using "linear" synthesis in very limited number of cycles without "denaturing". Do you have any experience for linear DNA synthesis, especially about the the...

11 December 2014 5,867 0 View

Which human genes are strongly suppressed by wild-type p53 in human cells?

I am interested in looking for those genes that are expressed more than five fold higher when p53 is lost or mutated.

11 December 2014 9,495 2 View

Which PCR product clean-up kit you would like to recommend?

I am trying to clean the PCR products for deep sequencing, and I would like to know which kit of which company you think is the best (based on your experience if you have done). Thanks.

11 December 2014 2,965 7 View

How likely DNA double strands are seperated (melted) at lower than melting temperature?

For example, how many percent (if any) of DNA is melted at 5 degree lower than the melting temperature? Thanks.

11 December 2014 578 8 View

Do you know of any high-yield genomic DNA extraction kit/protocol?

11 December 2014 945 3 View

Is digestion of genomic DNA helpful to increase the pairing of primer to a specific genomic region of interest?

We are trying to test the efficiency of pairing of a primer to a region of mouse genome. We are thinking of digesting genomic DNA before pairing and wonder whether the longer or the shorter...

11 December 2014 6,149 2 View

Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence?

I am on the lookout for the Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence. Does anyone know where I can find it? Thank you in advance

03 March 2021 3,568 1 View

In PRIMER, is it possible to downweight certain taxa in multivariate community datasets to reflect uncertainty around identification??

I have a dataset with about 80 different species. As usual, some species are very easy to identify with certainty whereas others are more difficult, which means that I am less certain of my...

03 March 2021 8,066 4 View

PACYC PCR not working after several attempts. Would anyone have any suggestions?

So, I have been trying to run a pACYC PCR which will be used later on for a Gibson Assembly. However the PCR is not working. I have already tried gradient PCR and changing extension time; however...

02 March 2021 1,146 2 View

Does anyone know HRM channel in a 2 plex QIAGEN Rotor- Gene Q qPCR can be used as a separate channel?

I am going to have 3 different probes in my qPCR work that I am going to do. But I realized that the machine we have in the lab is a Rotor-Gene Q 2plex HRM Platform, saying it has green, yellow,...

01 March 2021 8,544 1 View

Which cell line should we use to perform biological dosimetry experiments?

We are preparing some experiments based on irradiating cells under different conditions in order to evaluate the effects in terms of DNA damage, genetic expression, etc. As our project is...

01 March 2021 3,355 3 View

How to calculate the annealing temperature of a primer pair?

I have to amplify a gene and my primers just reached. The Tm for Forward primer is 64.2, and that of reverse primer is 65.5. Can some one suggest how to get the best annealing temperature? Thanks...

01 March 2021 360 7 View

Anybody with an idea on how do design specific primers for detecting tomato resistance genes tm-1, TM-2 and Tm2^2?

I am trying to identify these 3 genes among some tomato cultivar collections and after aligning some sequences from NCBI, I couldn't find unique sequences to target for specific primers. There...

28 February 2021 606 3 View

What is the ideal efficiency for a qPCR?

hello everyone, I need to do standard curves for my qPCR, what is the ideal efficiency range? I tried a primer (Mglu2 receptor) that gave an efficiency of 90.2%. Is it accepted?

28 February 2021 1,254 3 View

MiRNA qpcr problem?

Dear All, mirna primer showing some problem in the melting curve? any idea why? As attached is the melting curve. The forward sequence is obtained from miRBase and reverse primer is universal.

28 February 2021 5,008 4 View

Calculating volume of DNA required from DNA concentration?

Hi I am a bit confused. They are asking me to find out the volume of DNA required in ul (a total of 30-100 ng for genomic DNA) from the DNA concentration in the nanodrop reading which was 404.8...

26 February 2021 5,029 2 View