Using the histone H2a transcript as an endogenous standard to study relative transcript abundance during bovine early development.
The requirement for sample standardization is basic to any relative RNA abundance assessment. In 2002, we published on the RNA abundance profiling of several housekeeping targets during early bovine embryonic development. It was then concluded that histone H2a was the most stable transcript across the studied developmental period. Since that time, several teams have applied this information, yet neglected to use the published set of primers. Here, we show that these other primer sets do not target the same histone H2a variant. Within the present report, the RNA abundance profiles of their respective targets, for example, histone H2a.1, H2a.z, and H2a.o were measured in developmental series spanning the immature oocyte to the blastocyst stage embryo. In order to more clearly define the conditions that impact the RNA abundance level measurement of these candidates, the state of polyadenylation and the origin of the transcript either from de novo transcription or from maternal stocks were taken into consideration. The histone H2a.z transcript is principally regulated by de novo transcription following embryonic genome activation, whereas the levels of H2a.1 and H2a.o variants are largely of maternal origin, conferring a more stable profile throughout the studied developmental window. The histone H2a.z was found solely in a polyadenylated state whereas the H2a.1 and H2a.o were found to be more abundant in a nonadenylated form. The presence of serum in the in vitro embryo production system also had some impact on the histone H2a.1 RNA level at the blastocyst stage.
Vigneault
Christian
C
Centre de Recherche en Biologie de la Reproduction, Département des Sciences Animales, Université Laval, Québec, Canada.
Gilbert
Isabelle
I
Sirard
Marc-André
MA
Robert
Claude
C
D000818:Animals; D001483:Base Sequence; D002417:Cattle / Q000196:embryology* / Q000235:genetics; D046149:Embryo Culture Techniques; D047108:Embryonic Development / Q000235:genetics; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D006657:Histones / Q000235:genetics*; D008969:Molecular Sequence Data; D010802:Phylogeny; D020033:Protein Isoforms / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis*; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000592:standards*; D012689:Sequence Homology, Nucleic Acid
D006657:Histones; D020033:Protein Isoforms; D012333:RNA, Messenger