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Questions related from Michael Sacher
We are new to Gibson assembly and are using the NEB Gibson assembly master mix. Reactions with vector alone (no insert) give no colonies. Adding an inset of 660bp gave 10s of colonies. But a large...
24 July 2020 6,331 2 View
Is there a protocol available for freezing a human skin punch sample that can then be shipped and used to generate a primary fibroblast culture? Can someone send or refer me to the entire...
24 July 2018 4,286 1 View
How easy is this to do for someone who has never done this before? We have plenty of experience culturing cells but how easy is it to recognize when it has worked? And does anyone have a simple,...
09 May 2018 3,943 3 View
We are studying a gene with 5 exons. In a patient with a mutation in one of the introns we detect a splicing defect such that exon 3 is missing (exons 1,2,4,5). Yet we also detect the wild type...
14 March 2017 5,550 6 View
We would like to knock out several genes (independently) using the CRISPR/Cas9 system. We tend to use HeLa cells for our work for their ease of transfection for siRNA and for microscopy. But I am...
20 January 2016 7,773 7 View
It seems reasonable to me that a block in intra-Golgi traffic would affect ER-to-Golgi traffic. But I cannot seem to find a reference for this. Does anyone know of such a reference or have any...
30 November 2015 8,438 1 View
I cannot find a reference for this but assume it ot be the case. If ribosomes are bound to the rough ER during the translocation process, it stands to reason that upon inhibition of translation a...
22 October 2015 5,853 3 View
I understand that rRNA is made in the nucleus, but the ribosomal proteins are made in the cytosol following export of the mRNA from the nucleus. Since each nuclear transport step requires energy,...
31 May 2015 5,145 5 View
We suspect mycoplasma contamination in one of our cultures since DAPI staining reveals cytoplasmic staining. But one person suggested it was simply mitochondrial. This got me thinking: in all...
27 November 2014 2,374 18 View
Can a retroviral insertion in the intron of a gene affect the expression of that gene or affect promoter activity? What would the mechanism be? I imagine nonsense mediated decay would degrade the...
25 March 2014 5,323 1 View
We have found that when we boil our sample for 2 minutes in SDS PAGE sample buffer we can detect phosphorylated forms of our protein of interest. But when boiled longer (5 minutes or more), we...
30 September 2013 6,321 3 View
We have a large yeast strain collection and a plasmid collection. What software are people using to keep their collections organized and searchable? We want to be able to search auxotrophic...
12 August 2013 8,957 3 View
In some mutants we have looked at we see a defect in Ape1 processing under non-starvation conditions but a very mild detect under nitrogen starvation conditions. We interpreted that as our gene of...
01 May 2013 6,861 4 View
