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Questions related to Quantitative RT-PCR
I want to quantify the quantity of bifidobacterium in a milk sample. I have extracted DNA from bifidobacteria to be used as my standard, i did a 10-fold dilution and ran my standards with my...
04 April 2019 8,947 3 View
i'm trying to isolate RNA from primary osteocytes. I was able to obtain RNA samples with good purity until recently. I have tried the ethanol precipitation method with sodium acetate to purify the...
27 March 2019 978 4 View
Hello, I am doing DNA-SIP for 13C labeling soil samples lately and I checked the SCI papers which have done DNA-SIP work. Generally, after ultracentrifugation, we do density gradient fractionation...
18 March 2019 3,490 2 View
I'm performing RT-PCR analyses using actin, 36b4 and GAPDH as housekeeping. The samples are all concentrate around 10 ng/µL and the ratios 260/280 and 230/260 are in the range. Both controls and...
01 March 2019 8,621 3 View
I am working with embryos and oocytes, i am confused about how many an which internal controls to use for qPCR? in literature, i have seen only one internal control has been used and study is...
01 March 2019 5,710 1 View
I'm interested in using the Allen Brain Atlas API to download the coronal and saggital planes to map onto brain areas. I've previously been using the Paxinos 2007 atlas but would like to try...
21 February 2019 4,020 0 View
I used to think that signal peptides were necessary for a membrane protein to be displayed on the cell surface till I found that a GPR with multiple transmembrane structures have not a signal...
31 January 2019 1,275 3 View
I am working on sheep mRNA, and designing primers using NCBI gene and PrimerBLAST. For some genes, there are mRNAs under "RefSeq maintained independently of annonated genome" and also under...
23 January 2019 2,978 0 View
e but obviously i cant add the ALL PLATES as one of them was run at 10 mins holding time then after 7 mins the machine stopped due to emergency then i opened it again after a min and iu sat the...
19 January 2019 4,807 2 View
Using a mouse model, I will be collecting atrial and ventricular tissues at various ages. I will block the tissues in gelatin and generate 10um sections for immunohistochemistry. Does anyone...
03 January 2019 3,025 5 View
I am studying about how to do normalization. At first, I think that people have to calculate the log2 ratio as a primary step to convert the T=R/G ratio in microarray into another simple number to...
18 December 2018 2,733 2 View
I use 15% Sodium Carbonate solution for my research. When I wash the glassware with Acetone, a white precipitate would form on the wall. Any idea what that might be?
29 November 2018 4,610 3 View
Dear colleagues, I need support with my work in which I use the qPCR method. In general, in my experiments, I try to measure the level of induction of prophages in different culture conditions (...
22 November 2018 8,107 1 View
I conduct qPCR to determine the number of genome-containing particles in an AAV preparation using SYBR green technology. We used AAV-GFP plasmid as standard, and make 5 serial dilution (1:5) and...
21 November 2018 5,901 8 View
For example, if I have Ct = 18 for GAPDH and Ct = 20 for the gene of interest, is it possible to conclude that there are 4 times more transcripts of GAPDH than transcripts of the gene of interest?
21 November 2018 8,305 4 View
Hi, I engineered the yeast Saccharomyces cerevisiae to over-express some of its native genes, in addition to native copy of gene. For overexpression, I used strong modified GAL1 promoter ( that...
13 November 2018 608 3 View
I am searching for novel interacting Proteins through Yeast Two Hybrid System, using Matchmaker Gold Yeast Two -Hybrid System of Clontech. I have done all the experiments accordingly. At first...
08 November 2018 237 7 View
When performing qPCR, I have noticed that cDNA made from RNA preparations treated with Turbo DNase (Invitrogen) prior to reverse transcription produce CT values that are shifted several cycles...
01 November 2018 9,822 5 View
Dear all, I am quantifying bacteria via qPCR using SYBR-green assays and targeting the 23S rRNA-gene which is available in multiple copies in bacteria. My question is how do I accurately...
18 October 2018 7,803 2 View
i run qpcr for gen ki67 with reference gene beta actin. before do qpcr., I already optimized the annealing temperature in pcr. in gel electrophoresis there is no indication of dimer. but...
16 October 2018 4,729 3 View
Can anyone tell me primer sequences used for qPCR to quantify serotype DenV2 and DenV3? Thanks!
11 October 2018 8,053 2 View
I'm planning on exposing epithelial cells to particulate matter, and measuring cell death/viability. I will be using LDH assay on the supernatant, but for a second readout, I would like to use...
24 September 2018 4,908 4 View
Presently I'm using Lanciotti et al., protocol for detection of Dengue using SYBR green1 PCR and I need to extend it for quantification of copy number / viral load in patient serum samples. I...
20 September 2018 8,866 1 View
Hi, I'm doing doing RT-PCR to detect the presence (or not) of a pathogen. I've done it 6 six without any problems, however, my last three attempts failed, because I don't get a standard curve...
16 September 2018 4,055 6 View