The primers used for amplification of the AID transcripts were the following: forward 5'-CACCACTATGGACAGCCTCTTG-3' and reverse 5'-ACTGTCACGCCTC-TTCACTACG-3', and for endogenous control β-actin: forward 5'-CTGGCACCACACCTTCTACAA-3' and reverse 5'-CAGCCTGGATAGCAACGTACA-3'.
For amplicon sequencing, AID transcripts were PCR amplified using specific primers containing 5′-sequence tags.
From these results, we conclude that alternative AID transcripts AID-ivs3 and AID-ΔE4, though present at RNA level, were not detectable as translated protein at equivalent stoichiometry.
First, AID transcripts may be stored in the oocyte for translation after fertilization, which could associate this gene to a maternal-effect.
Concomitantly, a recent report showed that CLL samples expressing AID transcripts harbored significantly more γ-H2AX foci than non-AID-expressing samples when dsDNA repair by homologous recombination is inhibited, indicating AID-induced DNA damage [46].
AID transcripts were found in cells other than lymphocytes, most notably in mouse primordial germ cells and unfertilized oocytes (Morgan et al. 2004), and in testicular tissue from human biopsies (Schreck et al. 2006).
In our laboratory, we first reported that in contrast to normal circulating B lymphocytes, which only express AID transcripts following CD40L or LPS stimulation, in most CLL cases expressing UM VH genes, tumoral cells are able to express high levels of an active AID enzyme, although this expression is confined to a small proportion of the CLL clone.
To test whether AID transcripts in MEC1AID-SRC cells are not translated or otherwise immunoblot technique is not capable of detecting low-abundant proteins, we titrated different ratios of lysates of HEK293FT cells transiently transfected with respective GFP-AID cDNA constructs and performed immunoblotting.
As jurors and lawyers, aided by transcripts, listened to the tapes on earphones, Mr. Gotti passed up the opportunity to listen in or to read the transcripts.
This difference is not surprising because known genes are likely, on average, to be expressed at a higher level than novel transcripts, aiding their initial identification [ 36, 37].
To aid accurate transcript annotation, the NCBI pipeline incorporated RNA-seq data from a sooty mangabey pooled tissue reference sample, and data from 14 separate tissues produced through a joint effort by the Nonhuman Primate Reference Transcriptome Resource (NHPRTR; http://www.nhprtr.org/ 27 and the Human Genome Sequencing Center HGSCC) of Baylor College of Medicine.
