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Despite high similarity in sequence and catalytic properties, the L-lactate dehydrogenases (LDH) in lactic acid bacteria (LAB) display differences in their regulation which may arise from their adaptation to different habitats. We combined experimental and computational approaches to investigate the effects of fructose-1,6-bisphosphate (FBP), phosphate (Pi) and ionic strength (NaCl concentration) on 6 LDHs from 4 LABs studied at pH 6 and pH 7. We find: (1) The extent of activation by FBP (Kact)
S.pneumoniae D39 cells (wild type and delta greA) were grown in C+Y medium and cells were harvested for total RNA isolation at mid-exponential growth (OD600nm 0.3 for wt, 0.25 for delta greA). Total RNA was isolated as described before (Kloosterman et al 2006).
The total RNA samples were examined by capillary electrophoresis.
dephosphorylated with antarctic phosphatase followed by treatment with polynucleotide kinase (PNK).
Afterwards, samples were poly(A)-tailed using poly(A) polymerase. Then a
Stranded RNAseq libraries were prepared from 1µg total RNA from liver tissue using TruSeq Stranded mRNA library preparation kit (Illumina, San Diego, USA) using double unique indices (#20022371), according to the manufacturer's instruction (Part 15031057 Rev.E). Libraries were sequenced at the Norwegian Sequencing Centre (NSC). All libraries were pooled, and the same pool was sequenced on 4 flow cell lanes on a HiSeq 3000 machine (Illumina), generating 100bp single-end reads.
RNA sequencing files
RNAseq is utilised to validate the SnpEff annotation predictions for aberrant splicing. For this purpose the percentage exon retention for exons 4, 6 and 7 are calculated using RNAseq data.
RobOKoD algorithm was, designed then implemented as part of a study in RobOKoD: microbial strain design for (over)production of target compounds. (http://fairdomhub.org/publications/236). It was used to generate a strain of e.coli for producing butanol, that was then compared to an experimental strain. It was shown to perform better than similar methods (OptKnock, and RobustKnock).
RobustKnock algorithm was used as part of a study in RobOKoD: microbial strain design for (over)production of target compounds. (http://fairdomhub.org/publications/236). It was used to generate a strain of e.coli for producing butanol, that was then compared to an experimental strain.
A example of an RT-PCR Excel template. RT-PCR is Reverse Transcriptase PCR (NOT to be confused with Real Time PCR, which is normally referred to as qPCR)
This template was taken from the GEO website (http://www.ncbi.nlm.nih.gov/geo/info/spreadsheet.html) and modified to conform to the SysMO-JERM (Just enough Results Model) for transcriptomics.
Using templates will mean easier submission to public databases on publication and greater consistency of data in SEEK.