Welcome to "Science with J"

👋Hello world,

I present to you this blog where you can find out about scientific tips that will allow you to shorten your learning time in the analysis of scientific data.
We will also manually optimize basic bioinformatics protocols and provide bibliographic searching and filtering guidelines that can help you in Biomedical research.

In addition, we will inform you about advanced methods in the management of mathematical data related to the field of Molecular Biology, as well as image data related to the field of Cell Biology.


J.

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Primer validation/efficiency by qRT-PCR in five simple steps

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You have now amplified and measured the Ct of your serial cDNA dilutions... Now what? 😅 To measure the efficiency of your primer couple, you have probably been recommended the serial dilution method💧 🌡️ This method involves making dilutions of your typical control sample (e.g., 1 in 4 , 1 in 16 , 1 in 64 , 1 in 256 , 1 in 1024 ) and mixing it with your typical Master mix and your favorite primer concentration. This way you can confirm in silico whether the amplification pattern of your primers in successively diluted samples fits a trend line. How to measure the efficiency of your primer couple using the serial dilution method💦 1) If you measured Ct values ​​from at least 3 replicates of each dilution, calculate the Ct average of the 3 replicates for each dilution . 2) Consider your 1 in 4  sample as a hypothetical cDNA concentration value of 100 (100%), since that will be the highest sample concentration point. The remainder will be: 1 in 16 (25%), 1 in 64 (6.25%), 1 in 256 ...
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Design of expression plasmids using tools from custom vector suppliers

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🧬 Do you need an expression plasmid that is not commercially available (e.g. a mutated variant)? 😏 Whether you want the plasmid to contain a mutated gene or a wild-type coding sequence, here is a quick step-by-step guide.📋 The first thing is to find the coding sequence of the wild gene that expresses the protein you want. To do this, go to  https://www.ncbi.nlm.nih.gov/ and search the name of your interest gene using the Nucleotide search. Then click on the desired organism and select  mRNA . Choose the best entry for your query according to your requirement, and select the CDS  (and the protein sequence will appear on the right, a.k.a., translation ) .   Next,  copy-paste both FASTA text codes into a Word file . We will need them later. 👉If you only want to express the wild-type protein... Paste the CDS directly into the corresponding ORF item from the supplier (e.g.  VectorBuilder ).  https://en.vectorbuilder.com/design/pRP_Exp.html You wil...
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