Plant Molecular Biology ReporterThe Bxb1 recombinase mediates site-specific deletion in transgenic wheat Ann Blechl1, Jeanie Lin1, Min Shao2, Roger Thilmony1, James Thomson1*1 USDA-WRRC-ARS Crop Improvement and Utilization, Albany CA 947102 UC Davis Department of Plant Sciences, Davis CA 95616*Correspondence: James.Thomson@ars.usda.gov

Online Resource 1 The BxbI monocot codon optimized gene sequence (BxbNom) with its translation (using the single letter amino acid code) shown above the nucleotide sequence. The start and stop codons are underlined, and the added C-terminal nuclear localization signal is highlighted in yellow.

Online Resource 2 Comparison between a) growth chamber and b) greenhouse grown leaves from several T1 transgenic wheat plants. GFP activity (not shown) could readily be visualized in the tissues the day following bombardment, and GUS activity was present in the same cells after staining the tissues overnight at 37°C (a total of 2 days post bombardment before clearing the leaf tissue with ethanol).

Bxb145-35-2 Bxb145-35-4

Bxb145-35-3Bxb146-35-1

a) Leaves from growth chamber grown plants

Bxb145-35-7

Bxb145-35-2Bxb146-35-1 Bxb145-35-3 Bxb145-35-4 Bxb145-35-7

b) Leaves from greenhouse grown plants

pAHC15 (Ubi-GUS) pGUNG-BxbBPa

Online resource 3 β-glucuronidase enzyme activity detected in wild type ‘Bobwhite’ bombarded with pAHC15 or pGUNG-BxbBP. a) embryos b) ventral (crease) and dorsal (embryo) sides of endosperm. Leaky expression of pGUNG-BxbBP is noted in both embryos and the dorsal sides of endosperm.

BobwhitepAHC15

BobwhitepGUNG-BxbBP

BobwhitepGUNG-BxbBP

Ventral side Dorsal side

b