21 Questions 45 Answers 0 Followers
Questions related from Lyra Chang
Hi All, I have used MBP tag to help solubilize my target protein. It works really well. However, whenever I cleave off the His-MBP tag and try to clean up the sample by passing through another Ni...
09 March 2019 5,582 5 View
Hi All, We are planning to transfect a relatively large luciferase vector into cancer stem cells. Does anyone know now what would be the best transfection reagent for this purpose? Would...
11 September 2018 5,267 4 View
Hi All, I am trying to establish LC-MS based method to determine the methylation level of genomic DNA. I am currently using the DNA degradase Plus from Zymo research, however, there are usually...
18 June 2018 8,654 4 View
Hi All, Recently, I have finished my fragment screen and need to get protein-ligand co-crystal structures in order to guide the medicinal chemistry to improve the fragment hits I found in my...
17 February 2018 6,906 11 View
Hi All, We have a Thermo Scientific Multidrop™ Combi Reagent Dispenser and one of the tip of its small volume metal tip dispensing cassette was clogged so that the liquid cannot shoot straight...
13 November 2017 9,703 1 View
Hi All, I want to select a cell line for our drug study but it has to be highly sensitive to p53 induction - i.e. overexpression of p53 can easily kill the cell line. Any suggestion will be...
27 October 2017 1,079 2 View
Hi All, I need suggestion for good downstream target genes of cJun (or when I treat my cells with JNK inhibitors). I tested Notch1, FGF2, CCND1, FZD1, FRAT1, HDAC1 and BMP3 as downstream genes,...
12 May 2017 2,548 4 View
Hi All, I am doing drug discovery work related to DNA methylation level in cancer cells. I wonder if there are new & good assay that allow me to efficiently track DNA methylation status on a...
14 March 2017 5,894 11 View
Hi All, I have a bit of headache to find good downstream target gene to track for NFkB and Myc pathway activity. What I did was treating A549 cell with inhibitors of NFkB (BAY 11-7082) and Myc...
30 January 2017 6,185 4 View
Hi All, I was told that it is best practice to attempt growing protein crystals only after gel filtration column, but my protein is already quite pure after ion exchange column. I wonder have...
27 January 2017 3,552 6 View
Hi All, I am new to the FP assay. I am wondering if my protein is stored in 5% glycerol, would the FP result affected by the glycerol in the solution? Would glycerol reduce the tumbling of...
16 August 2016 8,289 6 View
Hi All, We have problem getting good transfection of PX459 by Lipofectamine 3000 into A549 cells (I am not 100% sure, just that we don't get any editing with all 4 gRNA we designed based on...
10 August 2016 7,593 8 View
Hi All, I have done tons of MTT assay lately for my project and have so many plates filled with the purple formazan solution (I used the SDS lysis method so no DMSO present). I just have no idea...
09 August 2016 10,106 2 View
Hi All, I am currently starting to develop a high throughput assay against my target which is a Ring E3 ligase. Does anyone have any successful experiences to share? Also, any input on good...
21 April 2016 8,471 2 View
Hi All, Our lab will have some money to buy equipment pretty soon and we are interested in buying a liquid nitrogen mammalian cell line storage chamber/freezer. For me, taking out frozen cells...
08 April 2016 1,158 6 View
Hi All, Since our lab doesn't have BL2 tissue culture room, I used other lab's space for generating Lentivirus and infecting the cells with virus. Now I am about to start my Puromycin selection. I...
11 March 2016 7,912 2 View
Hi All, I have done a first run of infection with several lentivirus containing different shRNA this week. I am about to add puromycin but just realized that I forgot to add polybrene! I will...
10 March 2016 578 12 View
Hi All, I am planning to use GIPZ Lentiviral shRNA to knockdown my target genes. I do know that those Lentivirus cannot replicate and its quite safe for us researchers. However, I have a 6 month...
22 January 2016 5,066 8 View
Hi All, I am planning to use 2 gRNAs and wt-CRISPR to KO my target protein from the cell lines I pick (ex. Hepatocellular carcinoma cell lines). I want to clone both gRNAs into vectors and...
25 December 2015 2,173 4 View
Hi All, I am new to the CRISPR-Cas9 system and wonder if there is any good protocol for cloning gRNA into vectors. Given that they are quite small in size, is there anything that I need to be...
25 December 2015 8,827 9 View
Hi All, Could anyone give me some suggestion as to what are the best assays for determine the oncogenic domain on a oncoprotein? Currently I am working on a protein that known to be upregulated...
13 July 2015 4,896 2 View
