Insect resistance. Hessian fly.
Contributed by Herbert Ohm and Lingrang KongHessian fly resistance gene H9 was recently re-located on chromosome arm 1AS (1, 2).
SCAR marker SOPO05909
This PCR marker was developed by Kong et al. (1). SOPO05909 maps at a distance of 2.8 cM from H9
SOPO05-F 5- CCC AGT CAC TCA TAT GCT ACC TAT -3 SOPO05-R 5- CCG AGT TGA TAT GCA CGA TG -3
PCR reaction mix
Total reaction volume: 25 µl
Liu et al. (2) determined that microsatellite loci Xbarc263 and Xcfa2153 flank H9 and H10 genes within a range of 0.3-0.5 cM. H11 is linked to flanking markers Xcfa2153 and Xbarc263 at 0.3 cM and 1.7 cM, respectively. The germplasm used as sources of Hessian fly resistance genes for this study were, Iris (H9), Ella (H9, ref. 10), Elva (H9 and H10), Joy (H10), Karen (H11).
PrimersBARC263-L 5'- GGA AGC GCG TCA GCA CTA GGC AAC -3' BARC263-R 5'- GGC TTC TAG GTG CTG CGG CTT TTG TC -3'Final concentrations of the products used in the PCR reaction:
PrimersCFD132-F: 5'- TTG TGC ATG ATG GCT TCA AT -3' CFD132-R: 5'- CCA ATC CTA ATG ATC CGC TG -3'Final concentrations of the products used in the PCR reaction:
For the H9 locus two RAPD markers are available H9-1 and H9-2. The first requires two primers (OPA09 and OPA17), while for the second only one primer is used (OPO05).
The reaction mixture was overlaid with 50 µl of light mineral oil if needed. Amplifications were carried out in a MJ Research PTC-100 thermocycler programmed for 40 cycles of 1 min at 94°C, 50 s at 36°C, 1 min at 72°C, and ending with 6 min at 72°C. The primers were used singly and in pairwise combination. The PCR products were fractionated either on 1.2% agarose gels in 0.5 × TBE buffer or by denaturing gradient-gel electrophoresis in 12% polyacrylamide with a denaturant gradient range of 10-50%.
Conditions presented here should be considered only as a starting point of the PCR optimization for individual laboratories.
1. Molecular mapping determines that Hessian fly resistance gene H9 is located on chromosome 1A of wheat. Kong L, Ohm HW, Cambron SE, Williams CE. In: Plant Breeding, 2005, 124, 525-531. DOI:10.1111/j.1439-0523.2005.01139.x
2. H9, H10, and H11 compose a cluster of Hessian fly-resistance genes in the distal gene-rich region of wheat chromosome 1AS. Liu XM, Fritz AK, Reese JC, Wilde GE, Gill BS, Chen MS. In: Theoretical and Applied Genetics, 2005, 110:1473-1480. DOI:10.1007/s00122-005-1982-z