To increase the transfection efficiency of naive CD8 T cells with Lonza Nucleofactor Kit?

I am working with Naive CD8 T cells and used LONZA Nucleofactor Transfection kit. The transfection efficiency with plasmids is quite less (30%). Does anyone have better experience with this or...

11 December 2017 8,766 3 View

How to type IFNAR-/- Mouse?

Does anyone know any successful method either PCR or FACS to type IFNAR-/- KO MOUSE?

05 June 2014 1,666 4 View

What statistical equation do I need to put in Standards results from BCA KIT (Thermo) to analyze my result?

The standards are not in a straight line and following a curve so how can I analyze the results accurately? Also I am using GRAPHPAD for my analysis.

04 May 2013 4,585 0 View

Thrombin activity in different buffers.

Is there any specific buffer for thrombin activity? I am currently using Tris-cl, pH 8.0 as cleavage buffer, but wish to use Na-phosphate buffer. Does this hamper the activity of thrombin?

01 February 2013 3,495 2 View

Purification of recombinant protein cloned in pET28c using buffer 7.0 by Ni-NTA Agarose.

I have a cell-free extract in K phosphate buffer pH 7.0 and I want to run Ni-NTA column to purify my protein. The protocol says to use buffers of pH 8.0. Is it necessary? Can we try a different...

08 September 2012 5,744 21 View

No title

03 March 2021 5,360 2 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

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

Has anyone used the Illumina ITS1 primer sets?

Hi everyone, Illumina provides a list of primers to amplify with high taxonomic coverage the ITS1 region for further fungal sequencing, but I cannot find the exact amount necessary of each...

25 February 2021 6,969 3 View

Genescan gives 3 peaks instead of one?

Hi all, I am doing a genescan analysis for a deleted exon in the patient sample. The amplified region is 215 bp. In the patient sample because of homozygous deletion, no peak is observed while in...

23 February 2021 5,645 1 View