Construction of transgenic plants overexpressing CS. Details of the
construct (A), genomic DNA PCR analysis (B) used in transformation of canola
with At-mtCS. A, Diagrammatic representation of pACS121-Hm. The gene, At-mtCS,
is expressed under the control of a constitutive 35S promoter. The construct
has selectable markers for NPT II and HPT II genes. The two primer pairs used
in genomic DNA PCR amplification are indicated as arrows in the construct. B,
Genomic DNA PCR analysis of transformants confirming the presence of At-mtCS
transgene. Genomic DNA was isolated from WT and two independent transgenic
lines (CS1 and CS12) and was used as template for PCR with two primer pairs
indicated above. 35S-F/AtCS-R, Expected amplification size of 0.7 kb; 35S/HPT
II-R, expected amplification sizes of 0.3 and 2.8 kb. Positive (+) control
includes plasmid DNA (pACS121-Hm), and negative (-) control includes PCR
reactions with no DNA.