Which Human Reference Transcriptome for RNAseq?

I am currently conducting a RNAseq experiment of different human blood cell populations. I decided to go with Salmon for quantification of the samples as it seems to be one of the best algorithms...

18 February 2020 8,329 3 View

How to compare marker expression of cells with different autofluoresence?

I my experiment I want to assess changes in monocytic cells by flow cytometry after a specific treatment. For example after treatment I want to compare CD14 expression of treated cells vs. control...

19 December 2018 687 10 View

Cell-Based Bioassay | Any tips on increasing assay window?

Hi all, I wonder if there are any tips reg. increasing the assay window? We perform 4-fold dilutions for 10 points. Currently in our case, if the initial concentration is relatively high, the...

19 January 2016 2,406 3 View

Absolute quantification qPCR - alternative splicing problem. Can you lend your expertise?

I have a problem with absolute quantification and standard deviations. When I do my standard curve (with gBlocks) and run all my samples the standard deviation between triplicates are around 0,1...

15 July 2014 7,091 3 View

Which is a lucid standard comprehensive book to guide on psychological scale construction and its standardization?

02 March 2021 7,082 3 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

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

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

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View

PUC19 as a transfer plasmid for lentiviral delivery?

Hello, Is it possible to use pUC19 as a transfer vector to be packed in using the second generation viral particles packaging system( pMD2.G; psPAX2 plasmids)? As far as I understand it there is...

28 February 2021 4,868 2 View

Did anyone ever try using pLVX-TetOne Plasmid with a copGFP as a selection marker instead of Puromycin?

When using a lentiviral vector for inducible expression of genes using the Tet-On system. In the literature, I saw a widely used of pLVX-TetOne-Puro Plasmid, is there a reason why people prefer to...

25 February 2021 1,011 3 View

Why do the PCR bands from the cDNA templates (extraction from the cell after transfection) show different size from the band of the plasmid template?

After transfection with the plasmid ( linearized ) and subcloning of the cell lines, RNA was extracted from the cell and then reverse transcripted to cDNA. When PCR reactions were run to verify...

25 February 2021 5,712 3 View

Does anyone have a sequence for the human CD34 promoter?

I'm looking for the human CD34 promoter sequence to engineer into a vector construct. Can anyone recommend a good source for this sequence? Thanks very much, Laura

25 February 2021 8,205 1 View

How do I transfect a large plasmid into PC12 cells with nerve growth factor (NGF) receptor?

I have a large 16 kb plasmid, which I need to transfect into PC12 cells. Lipofectamin 2000 didn't work and with GFP alone the transfection rate is very low. I also tried the neon invitrogen...

25 February 2021 6,635 5 View