27 Questions 23 Answers 0 Followers
Questions related from Liyana Azmi
I'm assembling 3 PCR fragments with my vector pBAD and transforming them into dh5a. The second PCR fragment has 4 variations to it and the template for this PCR fragment comes from a plasmid. I've...
02 February 2019 5,769 3 View
Im specifically looking for an anti-mouse antibody for flagellin (antisera H7). I've checked papers and googled extensively but couldnt find it. Any ideas?
04 April 2017 2,220 2 View
I need a protocol to shear flagella off E.coli O157:H7. I have been using a protocol online that is basically resuspending an overnight culture in a syringe and then, spinning it down to acquire...
03 March 2017 6,709 2 View
I have a protein which I know is stable with the presence of co-factors (NAD+ and Fe2+). Recently i have been trying to purify it with the TCEP as a reducing agent since my protein is known to...
11 November 2016 265 2 View
Hello, I have a protein that self associates into multimers and I have confirmed that using AUC. I would like to find the seqeuence in the protein that is responsible for self-association. Does...
05 May 2016 3,259 3 View
I am purifying a protein (47 kDa) that is fused onto an MBP (42 kDa). The PIs for both proteins are 6.48 and 4.9 respectively. After purifying it using a dextrose column, I have buckets of fused...
05 May 2016 9,064 8 View
I'm purifying a protein that is fused onto an MBP tag (42.5 kDa). My protein size is roughly the similar size of the tag (47kDa). Together, they form a complex about 89 kDa. After a run through a...
04 April 2016 8,726 3 View
I have sequenced my sample and found some mutaions in the fliC gene. I have found that they have introduced some aa changes however, I would like to know if this changes the conformation into R or...
03 March 2016 7,538 1 View
I have .dat files from scatter that contain the Dmax of a protein and now I want to generate ab initio models. however I dont know how to generate .out files that are needed byy DAMMIF programs on...
10 October 2015 9,171 1 View
I'm using a new column (5 ml Ni NTA HiFliQ column from Generon) to purify my protein AdhE which is Histagged and sized 96 kDa. However, during purification, the alarm pressure keeps going off even...
08 August 2015 5,244 10 View
I am purifying AdhE (96 kDa) protein which is bifunctional and ontains 2 functional domains; (acetaldehyde dehydrogenase and Alcohol dehydrogenase). Before purification, I did a small scale...
08 August 2015 4,210 7 View
I am doing Chip-seq, but at the end of the experiment I have obtained a clear DNA background for my negative samples but absolutely no DNA from my tested samples. I think there might be something...
08 August 2014 9,649 8 View
I used imidazole in my reference buffer when running sedimentation velocity and equilibrium experiments on AUC. however, I cant find the imidazole in the buffer range composition when using...
06 June 2014 3,181 0 View
I need 0.225M Mes/bis-tris buffer to optimize my crystal growth.
06 June 2014 9,642 0 View
My protein stays in solution with a high imidazole content (250mM imidazole) however, when analysing the results, imidazole is not within the buffer range, or rather, the program cannot compute...
06 June 2014 2,866 3 View
I am very new in crystallography and I'm just reading on microseeding techniques. There are so many terms on crystallography that I don't understand and before I actually do seeing, I hope I can...
06 June 2014 9,026 3 View
I am purifying a 35 kDa protein, its temperamental and oddly, it crashes in low imidazole rather than in a higher concentration of imidazole. Anyways, I wanted to dialyse it for SAXS analysis at...
05 May 2014 5,075 11 View
I made ampicillin plates (1:1000 ampicillin dilution that is 400ul in 4 mls of LB broth) and grew my transformants (dh5 alpha + vector pT7-FLAG2 + DNA inserts)into the agar overnight at 37 celsius...
03 March 2014 9,050 10 View
I have a protein sequence which I want to see in 3D. I tried Pymol and Cn3D, but the protein files that they have don't have the sequence which I want to see. I have to predict the protein...
03 March 2014 4,652 22 View
I did my birnboim and doly prep and received multiple bands everytime. I compared the bands with samples using Qiagen miniprep kits and still I have multiple clean bands. When I added RNase to my...
02 February 2014 5,931 3 View
I did a Birnboim and Boly plasmid Prep for my plasmid and insert which are grown on BL21. After that, I digested them with my restriction enzymes and run the samples on the gel. What I got was a...
02 February 2014 2,228 3 View
My vector is pT7 FLAG2 sized 4.8kbp and my insert is roughly 2-3kbp. I grew them in BL21 and sub-cultured them overnight with ampicilin before extracting them. After that I digested them with my...
02 February 2014 8,893 15 View
I'm screening for my plasmids which are pT7-FLAG2 with inserts approximately 1000-3000 bp each. My plasmid is 4.8kbp. I've cultured them in BL21 and tried extracting them using Birnboim and Doly...
02 February 2014 10,039 3 View
I am trying to express a 48 kDa protein in a pEt28a expresison vector. Attached is the map of the construct. I have tried expressing my protein in small aliquots just to see if my expression works...
01 January 1970 6,777 6 View
Roche provides an intereactive complete metabolic pathway map online however, I can't seem to be able to search for the metabolie/enzyme/anything on the map using the options provided. (here's the...
01 January 1970 2,567 2 View
Specifically, I'm looking for anti-FliD (also known as HAP2) for Escherichia coli O157:H7, the capping protein at the end of the filament.
01 January 1970 1,088 2 View
I'm purifying a protein that has a functional cysteine residue at the active site. In the file below, on the left is the SDS of the protein post-IMAC purification. These fractions are then...
01 January 1970 8,074 4 View
