How to know if my primers are suitable to clone a gene in pET28a vector to get histidine tagged protein?

08 August 2023 1 8K Report

Is there any online tool available to check, if designed primers are suitable to get overexpression of the histidine-tagged protein, cloned in pET28a vector?

How to confirm the suitability of primers for the same.

Usha Kantiwal

There are a few rules to design primers for gene cloning. If you follow them, there will be no issue. The rules can be easily get from PubMed. For in silico gene cloning, you can use the software SnapGene and ApE plasmid editor.

Badges
Science topic

More Nisha Rathor's questions See All

How does fluent FWH solve acoustic data at a location where no mesh is present?

In the simulation tutorial of the aeroacoustics around a cylinder by ANSYS, recievers are placed at a location where mesh isn't present, how does the solver predict the acoustics at that location...

16 October 2020 3,368 5 View

Problem with Cryosections?

Hi, I have a problem with cryosections while staining. Positively charged slides are used for cryosections, in spite of that during the staining process, the sections are washed away during the...

10 September 2020 3,767 3 View

Can we use 2, 6-dichlorophenol indophenol (DCPIP) method for analyzing crude oil degradation containing high sulfur content in the crude oil?

I was trying to find out whether my bacteria can degrade crude oil or not using DCPIP method that changes color after oxidizing hydrocarbons. But, I happen to order the crude oil containing 5%...

25 June 2020 9,801 2 View

Why my lenticrisprv2 plasmid after ligation of oligos is of smaller size than the expected?

after miniprep, I sequenced the plasmid. After sequencing I found that a region of plasmid was missing and its size if of around 8kb while it should be of around 12kb. It has happened to me for...

21 March 2020 736 3 View

HnE Protocol for the frozen section of cells encapsulated in hydrogels?

Hi all, I'm facing problems in staining the cryosection of the cells encapsulated in hydrogels. I'm using both charged slides as well as the albumin coated slides for the HnE staining. During the...

28 May 2019 1,197 5 View

VZV plaque assays?

Has anyone done plaque assays for VZV using toluidine stain?

10 March 2019 7,657 1 View

Has anyone done co-transfection with siRNA and the fucci system viral vectors by Thermofisher (Cat: P36238) ?

I'm trying to knockdown my protein in adherent cells and label cells with the fucci system for cell-cycle segmentation using microscopy. My protocol is as follows Day 0 : siRNA...

07 August 2018 4,362 4 View

Can anyone suggest me some ph.d research topic from machine learning and deep learning?

I am currently pursuing Ph.d

01 January 2018 7,222 17 View

What is the procedure for performing ELISA on protein pellet extracted from cell lysate?

Hi all, I wanted to perform ELISA (sandwich elisa) on the protein pellet extracted from cell lysate. I would like to know in which buffer, the protein pellet should be resuspended? Thank you Nisha

10 December 2017 9,325 3 View

Acceleration time history for Korba earthquake in India?

Can I get it in matlab format or notepad

08 October 2017 7,788 3 View

Can linear DNA be used as template for site-directed mutagenesis?

Is it possible to induce site-directed substitution mutation by quick-change method on linear dsDNA? or it has to be cloned in some vector? If yes, should it be treated with the Dpn1 enzyme...

03 March 2021 401 4 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 have a recommendation for yeast reporter gene plasmid/protocol?

Hi, could anyone recommend a plasmid and/or protocol for reporter gene assay in S. cerevisiae? I want to assess the effect of growth conditions on a transcription factor, so I want to clone it`s...

01 March 2021 210 1 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

Why might my GFP tag be dissociating from my plasmid after transfection?

I have a set of stably transfected cell lines all transfected with plasmids containing GFP tags on the C terminus. During a western blot using anti-GFP antibody, one of my plasmids has dissociated...

01 March 2021 9,310 4 View

Precipitate after adding laemmli buffer to protein solution containing Potassium Acetate?

Hi, I am running a size exclusion chromatography experiment with a buffer containing Potassium Acetate as a salt. I analyse these fractions through SDS-PAGE. After boiling my SEC fractions in...

01 March 2021 2,622 3 View

Expression cloning by using cDNA,do I need to modify the cDNA first?

I am going to have a expression cloning of mammalian gene by using shuttle plasmid to transforming the E.coli However I don't know I should only inserting the Coding sequence ,or I can...

28 February 2021 5,440 3 View

What is the most suitable server for predicting protein structures in bacteria?

I have used the i-Tasser several times, however it has been unavailable for several days. I tried the swiss-model, but the output was not very pleasant due to the model used. Is there any other...

28 February 2021 4,521 3 View

Do you need the CMV enhancer before the CMV promotor for a proper function of CMV on the gene that follows?

I'm a student and I have to produce a cell line with a knockin for the NRF2 gene with GFP. I have to put a promotor in front of the GFP because the gene will be too far away from the promotor of...

28 February 2021 7,127 2 View