To obtain insertion alleles in AML1 and other AML gene family members, we screened the insertion allele populations at the University of Wisconsin Arabidopsis Knockout facility [ 25], obtaining alleles harboring insertions in AML1, AML3, AML4 and AML5 (Fig. 3).
This group analysed single and multiple mutants of the eight ACS genes in Arabidopsis using a combination of T-DNA insertion alleles and amiRNA technology.
Using semi-quantitative and qRT-PCR, we demonstrate that these fln insertion alleles do not produce detectable mRNA and are null alleles, and through complementation and characterization of multiple alleles, indicate that the observed phenotypes result from loss of fln expression.
Additionally, transposon insertion alleles can be uniquely informative: a single insertion may be sufficient to generate a gene disruption, reporter gene fusion, epitope-tagged allele, and conditional mutation.
Until now however, due to the inherent size difference associated with insertion and no insertion alleles, the use of Alu REs has not been practical for forensic applications.
Three independent T-DNA insertion alleles (SAIL_61_B05, SAIL_390_D05, SALK_108979) were obtained from the Nottingham Arabidopsis Stock Centre NASCC).
Two pde166 (alb1) insertion alleles from the Syngenta collection [6] are included in the SeedGenes database.
The primer pair of LBa1 and RP specifically amplifies DNA from the insertion alleles, and the primer pair of LP and RP was used for identification of the non-insertion alleles.
Similar phenotypes were observed with two additional P element insertion alleles captk01217 and capt06955 (data not shown).
However, no insertion alleles were identified.
