What are the primers to get whole length of the sequence?

Dear all Please if someone can guide me.. I have cloned a gene of 1500 bp pairs in FAST-Plasmid. I used reverse primers to sequence the gene. The company sent me 900bp length.. I want to sequence...

11 December 2015 8,729 1 View

Anyone familiar with restriction double digestion of p35S-FAST Plasmid?

10 November 2015 7,135 2 View

Which (forward or reverse, or both) primers should I use for cDNA synthesis from gene specific primers?

I have extracted RNA from plant immature fruit. I want to synthesize its cDNA by gene specific primers. Please guide me on which (forward or reverse or both) primers are used in such cases.

03 April 2014 9,362 6 View

Can you help me achieve adequate amplification of required band?

I am using 0.2mM dNTPs,40pm primers,0.6microlitre Taq,2 microlitre MgCl2(2.5mM) in a total 20 microlitre reaction volume for PCR. I have kept the temperature the same so far. I need a band size of...

02 March 2014 1,460 13 View

Would the process of extracting RNA from plants without liquid nitrogen be successful?

Would the process of extracting RNA from plants without liquid nitrogen be successful?

02 March 2014 4,430 9 View

Would RNA extraction from plants be successful if i do not use Liquid nitrogen?

Thanks in advance

02 March 2014 3,995 1 View

How should my cDNA band look like on gel?

I want to synthesize cDNA from plant RNA. Please tell me that what should the cDNA band look like on gel, so that I can guess that cDNA has successfully synthesized. Please show me some gel...

01 February 2014 887 1 View

Can anyone help with RNA storage?

I stored plant RNA samples at -20 degree centigrade for 5 days and then I shifted to -80 degrees. Can I use this RNA for cDNA synthesis?

01 February 2014 6,004 7 View

Can anyone help with optimizing PCR conditions for my gene of interest?

I made different dilutions of control cDNA and in each case after running its PCR got the band of 496bp .The Gel pic is as below. But when I ran PCR for my gene of interest with same dilutions of...

01 February 2014 652 3 View

Can anyone help with cDNA agarose gel?

I have synthesised cDNA from plant RNA .The gel picture is given below. Please if anyone could tell me that whether the cDNA has synthesised or not? If synthesised, can I use this for Reverse...

01 February 2014 3,377 13 View

Buffer to disrupt platelet alpha granules for PF4 quantification?

Hello! I want to quantify by ELISA the secreted (from platelets poor plasma) and the non-secreted (from platelet lysate) PF4 before and after TRAP stimulation. I will use the ELISA from R&D...

03 March 2021 1,499 2 View

Buffer Exchange using column packed with sephadex G-25 effect on protein concentration?

Hello, If i am doing a buffer exchange for an antibody of 1mg/mL, does the elution lose protein in the process of buffer exchange? For example, if i flow through 500 uL of 1mg/mL sample, and...

03 March 2021 6,299 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

How optimize the Size Exclusion Chromatography?

Hello! I'll do a size exclusion chromatography, but I only have an open column, and I'll perform the peptide extraction from yeast, using buffer lysis (sodium phosphate 50 mM/NaCl 30 mM/DNAse and...

01 March 2021 2,215 2 View

Why are my bands weird in this Agarose gel?

Can someone please give me some possible things that could go wrong? Here is my recipe: 0.5g Agarose 50 mL of TAE 1x 1 uL ethyl bromide. Gel was run at 100V for 1 hour. The buffer used is also TAE.

01 March 2021 9,952 3 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

What do I do with DLS peaks that are from the buffer?

I'm looking at the aggregation of my protein sample using DLS. Unfortunately, my buffer (20mM HEPES) also results in a set of peaks. These are at approximately 1 and 1000 d.nm. The lowest peak...

01 March 2021 9,015 2 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

Which group formation theory do you recommend for projects that aim to bring together entrepreneurs for training for regional development?

I would like to have ideas about which theories of group formation are suitable for entrepreneurial training projects, with the aim of creating bonds, building trust, and collaboration for...

28 February 2021 4,223 2 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