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Questions related from Enes Yağız Akdaş
Dear All, I use HEK293-T cells for preperation. I treat them 24h with epoxomicin. And then I isolate proteins using RIPA buffer. I precipitate overnight 5 mg protein with 25 micro liter of...
30 March 2024 3,379 0 View
Dear All, I am encountering challenges with AAV-mediated neuronal transduction, where high volumes lead to significant cell death, while low volumes yield inadequate signal levels. Here's a brief...
23 March 2024 4,320 5 View
Dear All, I differentiate some new clones of hiPSCs into neuronal progenitors using dual SMAD inhibitor in DMEMf12/neurobasal mix+ NEAA +n2+b27. After induction of 12 days in culture and...
25 February 2023 616 1 View
Dear All, I am currently performing a stemcell differentiation into neuronal progenitors. However, I`ve realized that the incubator stopped heating yesterday so that my cells were kept under 23...
12 February 2022 7,566 3 View
Dear all, I do have human embryonic stem cells. I differentiate them into NPCs. However, when I perform qPCR for various genes, I also detect cT values for tuj1 in stemcels. In neurons, the...
07 March 2021 1,966 4 View
Dear all, For a paper, I need to show that my hESC can differentiate into 3 layer. I have neurons which represents for ectoderm. I need some cells ( really type is not matter) that represent...
22 October 2020 3,520 5 View
Dear all, I have flash-frozen mice brains for RNA sequencing and ATAC sequencing. However, in flash-frozen tissues isolating nuclei is problematic. And I can only use one side of the hippocampus,...
06 September 2020 4,700 0 View
Dear All, I am working with thermo fisher H9 cell line, human induced pluripotent stem cell. I made a mutation on them with crispr cas-9 I obtain single colonies and I propagate my clones. I...
14 June 2020 9,157 3 View
Dear all, I have antibiotics free stemcell culture. In the last passage, 2 days after I got bacteria contamination in all wells. I probably contaminated them during passaging because they were...
26 May 2020 7,706 0 View
Dear All, In AAV vector if we clone a miRNA we clone this pri-miRNA sequence in to the 5' of the reporter gene. For example this is the empty vector : https://www.addgene.org/21893/ and this...
25 May 2020 1,744 0 View
Dear all, I am trying to learn electrophysiology. And I have a problem that I didn't understand the reason. I am adding 3 picture belongs to my experiment during baseline recording with 20...
14 May 2020 2,967 0 View
Dear all, I have an RNA seq data and I have many genes whose expression are altered by my treatment. However, The up-regulation pattern is generally about %50 (which makes Log2foldChange value 1....
08 May 2020 1,333 7 View
Dear all, I cloned 25bp fragment into a vector. My no-insert ligation has no colonies and my insert+vector ligation has many. Which is the first clue that I have cloned the fragment. Than I...
23 December 2019 2,174 7 View
Dear friends, We kindly get U-251 MG cell line as a gift from an other lab. after 3 passage we have determined that cell line has yeast contamination. Each stock is contaminated while other...
10 December 2018 6,035 8 View
hey, I have generated stable cell lines with random entegration of pCMV-miR vector. Vector contains a neoR gene fused with GFP with IRES. I got some GFP+ and GFP- colonies. and GFP- colonies...
22 August 2018 6,146 3 View
I am working with glioblastoma U-87 cell lines. I have pCMV-miR expression vector ~8kb. I tired both Transfast, Fugene HD, PEI, Jetprime. They had max %30 efficiency. Transfast best with 1:1...
29 April 2018 1,222 7 View
Hey, I have a problem during my plasmid electrophoresis. I got my plasmid band in the same line with 15kb marker. But when I digest it with a restriction enzyme I got my band in the correct...
21 February 2018 5,342 6 View
Hi, I am trying to clone miR96 precursor into the pcmv-miR expression vector. These are my primers; GCGATCGC: SgfI ACGCGT: MluI Bold sequences are overhanging...
19 January 2018 7,809 2 View
Hey, I have clonned a double stranded oligo into glo-mir vector. My vector is 7.5kb and insertion is 81bp. I have isolated my plasmid from colonies after trsnsformation and perform agarose gel...
30 November 2017 7,899 3 View
Hey, I will clone a miRNA into the expression vector. I have 2 primer sets to get full lenght pre-miRNA, as it is shown belove; Set 1: 5' _____________3' Set 2: 3'...
24 October 2017 7,333 6 View
I wonder that , does the DNA polymerase deattach from the DNA double helix after the sequens between two primer is complated? What I am trying to ask is that, If I have a 200 nt single strand...
21 July 2017 9,197 3 View
I am working with a protein whose 3' UTR sequence is targeted by some of the miRNAs. I will perform luciferase reporter assay to be sure about that these miRNAs cut my mRNA's 3'UTR sequence. But...
17 July 2017 4,115 3 View
Dear friends; I have monoclonal CTL2 antibody. And it is known that CTL2 protein has only 2 isoforms thar are only ~5 aminoacid changed. Bu after 14h incubation at +4 with primary antibody, I get...
29 March 2017 7,817 8 View
I have an acid-phenol:chloroform for RNA isolation to remove DNA, but its pH isn't acidic anymore. Can I change the pH with HCL, or will I have to prepare it again?
08 February 2015 2,590 3 View
Dear All, In most protocols who maturate neurons from IPSC-derivered NPCs (neuronal progenitor cells); they use NEAA in stemcell stage and NPCs induction phase, some of them use NEAA in NPC...
01 January 1970 1,771 4 View
