Disease resistance. Septoria Tritici Blotch Resistance.

Stb 4

Microsatellite marker Xgwm111

Locus Xgwm111 is located on chromosome 7D, 0.7 cM from Stb4.

   WMS111-L   5'- TCT GTA GGC TCT CTC CGA CTG -3' 
   WMS111-R   5'- ACC TGA TCA GAT CCC ACT CG -3' 
   

• 36 ng genomic DNA
• 0.64 µM each primer
• 800 µM each dNTPs
• 0.04 U Hot Star Taq DNA polymerase
• 5.8 µl of sterile deionized water
• 2 µl 10x PCR buffer
• 1 mM MgCl₂

Total volume: 25 µl

• Denaturing step: 94°C, 3 min
• Amplification step (44 cycles):
• 94°C, 1 min
• 55°C, 1 min
• 72°C, 2 min
• Extension step: 72°C, 7 min

PCR products were separated by electrophoresis in 3% agarose gels. An approximately 210-bp DNA fragment is amplified from the resistant parent Tadinia (red arrowhead), compared to a band of approximately 220-bp in the susceptible parent Yecora Rojo (green arrowhead). A second locus segregates among these progeny, producing bands of approx. 150-bp, but is not linked Stb4.

Microsatellite locus Xgwm111. The photo shows DNA bands amplified from two parents and 8 F10 resistant (R) and susceptible (S) lines derived from a cross between wheat cultivar Tadinia (R) and the susceptible parent (Yecora Rojo × UC554). The bands were resolved on a 3% agarose gel. M: molecular size marker (25-bp ladder). The red arrowhead points to the band of the R genotype, and the green arrowhead signals the band position of the S genotype. The bands of around 150-bp belong to a second locus unrelated to Stb4.

Conditions presented here should be consider only as a starting point of the PCR optimization for individual laboratories.