Identification of genes from flat oyster Ostrea edulis as suitable housekeeping genes
for quantitative real time PCR.
In this study, the 152 strains were further analyzed by the reference methods, including molecular biological analysis (16S rRNA gene sequence analysis and housekeeping gene
sequence analysis), and physiological and biochemical analysis if necessary.
In this study, the reaction of housekeeping genes
(2004) Determination of stable housekeeping genes
, differentially regulated target genes and sample integrity: BestKeeper-Excel-based tool using pair-wise correlations Biotechnology Letter 26, 509-515.
To ensure the integrity of these results, the additional housekeeping gene
18S rRNA was used as an internal standard to demonstrate that ACTB and PBGD mRNAs were not differentially regulated in the groups of GCs.
[DELTA]Ct values were normalized using the mean values of three housekeeping genes
: [beta]-Actin, [beta]-2-microglobulin, and GAPDH.
The melting curves of the 2 housekeeping genes
ACTB and HTN3 showed a single peak with similar melting temperatures (85 [degrees]C), indicating no DNA contamination and no primer-dimer artifacts.
Based on earlier gene expression studies in domestic fowl, we have tested the stability of expression of seven housekeeping genes
, including GAPDH, ACTB, HPRT1, TUB, succinate dehydrogenase flavoprotein A (SDHA), 18S rRNA and 28S rRNA in this study.
PCR Amplification of the Housekeeping Genes
and Polymorphism Analysis
In Chapter 3, the authors cover topics such as the quality and quantity of the labeled target, the use of housekeeping genes
and other controls, imaging of the arrays, and initial statistical and data analysis steps.
To characterize the Leptospira species detected in bats from Mengyin County, we attempted multilocus sequence typing (MLST) on 7 housekeeping genes
(glmU, pntA, sucA, tpiA, pfkB, mreA, and caiB), as previously described (9).
Specific primers for determining the expression of housekeeping genes
and the genes of interest (Table 1) were designed using Primer 3 software.