Insect resistance. Greenbug.

Gb3 gene

Contributed by Yiqun Weng (y-weng@tamu.edu )and Jackie C. Rudd

Weng et al. (6) identified five microsatellite markers (BARC76, GWM037, GWM428, WMC634 and WMC824) that are closely linked with Gb3 based on agarose or polyacrylamide gel-based electrophoresis. Two AFLP markers, XMgccPagg and XMaggPatg, are also tightly linked with Gb3 in the population analyzed (5). More information on these markers can be found in Refs 5 and 6.

Marker Xwmc634

The Gb3 gene co-segregates with locus Xwmc634, and it was found to be polymorphic for parents with or without the greenbug resistance gene. Other microsatellite loci are located close to Gb3 and they can be used in case of lack of polymorphism (see map below).

   WMC634-F   5'- AGC GAG GAG GAT GCA TCT TAT T -3' 
   WMC634-R   5'- GAC ATA CAC ATG ATG GAC ACG G -3' 
   

Final concentrations of the reagents used in the PCR amplification 50-100 ng genomic DNA 0.5 µM of each primer 200 µM dNTPs 1 U Taq DNA polymerase 1x PCR buffer 1.5 mM MgCl2 Total volume: 25 µl PCR conditions: Denaturing step: 94°C, 3 min Amplification step (35 cycles): 94°C, 1 min 50°C, 1 min 72°C, 2 min Extension step: 72°C, 10 min

After PCR amplification, products were separated on 6% denaturing polyacrylamide gel. The Xwmc634 allele linked to Gb3 has a size of around 208-bp.

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