Transcript Use of the Roundup ReadyTM Trait to Estimate Selfing in

Use of the Roundup ReadyTM Trait
to Estimate Selfing in Hand Crosses
of Alfalfa
Mark McCaslin and Stephen Temple
Forage Genetics International
Background
• Alfalfa is predominately cross pollinated
• % selfing is difficult to determine without
simple and effective genetic markers
• Transgenic traits offer unique advantages in
studying alfalfa genetics and reproductive
biology.
– Cross vs self pollination
– Pollen flow studies
Advantages of the Roundup
Ready Trait
• CP4 gene inherited as a single gene dominant trait
• Easily identified phenotype
– Efficient high-throughput screening
• Multiple transgenic events available with eventspecific PCR primers.
Event specific PCR
Promoter
TP
CP4 Gene
Terminator
Experimental Design
• Two populations of plants (~50 plants each)
– Pop B – simplex plants containing transgenic
event B (Bxxx).
– Pop D – simplex plants containing transgenic
event D (Dxxx).
• Non-emasculated hand crosses were made in
greenhouse Pop D (female parent) x Pop B
Experimental Design
• Seed harvested from Pop D plants
• Progeny were sprayed with Roundup
• RR progeny were tested with eventspecific PCR to identify the RR
genotype
– BD dihomogenic
– D event only
– B event only
Results
• 74.4% of the progeny were tolerant to Roundup
(75% expected).
• Genotypes of the RR progeny were established
with event-specific PCR and frequency of each
genotype was compared to theoretical value.
Genotype
BD dihomo
# plants
2068
% actual
30.5
% expected
33.3
B only
D only
1923
2776
28.4
40.9
33.3
33.3
Discussion
• Increased frequency of D event only
progeny and decreased frequency of B
event only and BD dihomogenic is likely
due to unintentional selfing during hand
crossing.
• Data suggests ~10% selfing in this cross
• Similar results from other crosses over
multiple years, locations and genetic
backgrounds.
Discussion
• Other pollination related experiments
planned or in process:
– Cross vs self-fertility with leafcutter bees in
commercial seed production setting.
– Pollen flow studies with honeybees