Both sequencing and CAPS assay confirmed that all three predicted mutants were real.
Third, the epistatic interactions, which are deduced from single and double mutants, are incomplete because real epistasis underlying the growth traits could exist among more than two genes.
Nowadays, this is the best way of obtaining the "mutant"/changed phenotype for analysis of gene function and plant improvement, especially for species with large genomes, like cereals, for which real mutants are for most of the genes unavailable [ 1, 4, 12- 15].
These large deletions may be real biological mutants or they could be amplification or sequencing artifacts (e.g. chimeras).
The extent of gene silencing in the five putative knock out mutants was quantified by Real Time RT-PCR.
The New Mutants are the outcasts.
Fortunately, these mutants are rare.
Notably, significant down-regulation of mab21l2, rx2, hmx1, six6b, vsx2, aldh1a3 and rorb transcripts in 13 hpf rx3 -/- mutants was confirmed by real-time PCR.
2) There is real concern about expression levels in the mutants used.
