Fig. 3.

Arabidopsis eIFiso4G knockout lines. a Genomic screening. i4g1 lines carry a pROK2 T-DNA insertion in exon 9. i4g2 lines contain a pROK2 T-DNA insertion in exon 4. PCR screening of genomic DNA with gene specific primers yields a 928 bp band which is absent in homozygous i4g1 plants, but present in both wild-type Col-0 and heterozygous plants. Screening with a combination of pROK2 specific and gene specific primers yields a band of 601 bp in i4g1 and heterozygous plants that is absent in Col-0. PCR screening of genomic DNA with gene specific primers yields a 915 bp band which is absent in homozygous i4g2 plants, but present in both wild-type Col-0 and heterozygous plants. Screening with a combination of pROK2 specific and gene specific primers yields a band of 753 bp in i4g2 and heterozygous plants that is absent in Col-0. b Confirmation of eIFiso4G isoform knockout by RT–PCR and western blotting. mRNA populations from wild-type and mutant plants were probed for actin, eIFiso4G1, and eIFiso4G2 message (left panel). Western blotting of total plant extracts probed with antibody to AteIFiso4G1 (right panel upper). Coomassie stained Rubisco is shown for a total protein loading comparison (right panel lower). c Phenotypic effects. (Left panel) Overhead comparison of plants at 21 days post sowing. (Right panel) Profile of plant lines at ~56 days post sowing