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A DNA-based method for studying root responses to drought in field-grown wheat genotypes.

Huang CY, Kuchel H, Edwards J, Hall S, Parent B, Eckermann P - Sci Rep (2013)

Bottom Line: Root systems are critical for water and nutrient acquisition by crops.Current methods measuring root biomass and length are slow and labour-intensive for studying root responses to environmental stresses in the field.The new method eliminates the need for separation of roots from soil and permits large-scale phenotyping of root responses to drought or other environmental and disease stresses in the field.

View Article: PubMed Central - PubMed

Affiliation: Australian Centre for Plant Functional Genomics, The University of Adelaide, Waite Campus, Glen Osmond, South Australia, 5064, Australia.

ABSTRACT
Root systems are critical for water and nutrient acquisition by crops. Current methods measuring root biomass and length are slow and labour-intensive for studying root responses to environmental stresses in the field. Here, we report the development of a method that measures changes in the root DNA concentration in soil and detects root responses to drought in controlled environment and field trials. To allow comparison of soil DNA concentrations from different wheat genotypes, we also developed a procedure for correcting genotypic differences in the copy number of the target DNA sequence. The new method eliminates the need for separation of roots from soil and permits large-scale phenotyping of root responses to drought or other environmental and disease stresses in the field.

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Related in: MedlinePlus

Genetic variation in copy numbers of TaITS2 per genome among 20 wheat genotypes.Total root DNA was isolated from primary roots of one month-old plants grown in a potting mix, and used to determined copy numbers of TaITS2 per genome using quantitative real-time PCR. Ct values of TaITS2 and TaPHT1;6A using sequence-specific primers and probes were determined for all samples. The copy number of TaITS2 per genome was calculated by 2ΔCt (ΔCt = TaPHT1;6A Ct – TaITS2 Ct). Predicted means of four biological replicates are presented.
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f1: Genetic variation in copy numbers of TaITS2 per genome among 20 wheat genotypes.Total root DNA was isolated from primary roots of one month-old plants grown in a potting mix, and used to determined copy numbers of TaITS2 per genome using quantitative real-time PCR. Ct values of TaITS2 and TaPHT1;6A using sequence-specific primers and probes were determined for all samples. The copy number of TaITS2 per genome was calculated by 2ΔCt (ΔCt = TaPHT1;6A Ct – TaITS2 Ct). Predicted means of four biological replicates are presented.

Mentions: Genotypic differences in copy numbers of wheat target sequence ITS2 (TaITS2) among 20 modern Australian spring wheat genotypes were determined using qPCR of TaITS2 and TaPHT1;6A, a single-copy gene, (Supplementary Fig. 1). Ct values of TaITS2 and TaPHT1;6A from root DNA samples were used to calculate TaITS2 copy number per genome. The Ct values of TaPHT1;6A were similar among 20 genotypes (27.68 ± 0.06), whereas the Ct values of TaITS2 among 20 genotypes with sequence-specific primers, ranged from 13.82 ± 0.20 to 16.16 ± 0.18. The genotypic differences in the Ct value of TaITS2 were highly significant (P < 0.001). Calculated copy numbers of TaITS2 varied from approximately 2,900 copies per genome in the variety Seri to 15,000 copies per genome in Young (Fig. 1). The average copy number of TaITS2 in the 20 genotypes was approximately 8800 per genome, comparable to 8200 rDNA repeat units per genome estimated for the two major rDNA loci (Nor1 and Nor2) in wheat by DNA hybridization29. The results indicate that there is genotypic variation in TaITS2 copy numbers among the 20 wheat genotypes.


A DNA-based method for studying root responses to drought in field-grown wheat genotypes.

Huang CY, Kuchel H, Edwards J, Hall S, Parent B, Eckermann P - Sci Rep (2013)

Genetic variation in copy numbers of TaITS2 per genome among 20 wheat genotypes.Total root DNA was isolated from primary roots of one month-old plants grown in a potting mix, and used to determined copy numbers of TaITS2 per genome using quantitative real-time PCR. Ct values of TaITS2 and TaPHT1;6A using sequence-specific primers and probes were determined for all samples. The copy number of TaITS2 per genome was calculated by 2ΔCt (ΔCt = TaPHT1;6A Ct – TaITS2 Ct). Predicted means of four biological replicates are presented.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824167&req=5

f1: Genetic variation in copy numbers of TaITS2 per genome among 20 wheat genotypes.Total root DNA was isolated from primary roots of one month-old plants grown in a potting mix, and used to determined copy numbers of TaITS2 per genome using quantitative real-time PCR. Ct values of TaITS2 and TaPHT1;6A using sequence-specific primers and probes were determined for all samples. The copy number of TaITS2 per genome was calculated by 2ΔCt (ΔCt = TaPHT1;6A Ct – TaITS2 Ct). Predicted means of four biological replicates are presented.
Mentions: Genotypic differences in copy numbers of wheat target sequence ITS2 (TaITS2) among 20 modern Australian spring wheat genotypes were determined using qPCR of TaITS2 and TaPHT1;6A, a single-copy gene, (Supplementary Fig. 1). Ct values of TaITS2 and TaPHT1;6A from root DNA samples were used to calculate TaITS2 copy number per genome. The Ct values of TaPHT1;6A were similar among 20 genotypes (27.68 ± 0.06), whereas the Ct values of TaITS2 among 20 genotypes with sequence-specific primers, ranged from 13.82 ± 0.20 to 16.16 ± 0.18. The genotypic differences in the Ct value of TaITS2 were highly significant (P < 0.001). Calculated copy numbers of TaITS2 varied from approximately 2,900 copies per genome in the variety Seri to 15,000 copies per genome in Young (Fig. 1). The average copy number of TaITS2 in the 20 genotypes was approximately 8800 per genome, comparable to 8200 rDNA repeat units per genome estimated for the two major rDNA loci (Nor1 and Nor2) in wheat by DNA hybridization29. The results indicate that there is genotypic variation in TaITS2 copy numbers among the 20 wheat genotypes.

Bottom Line: Root systems are critical for water and nutrient acquisition by crops.Current methods measuring root biomass and length are slow and labour-intensive for studying root responses to environmental stresses in the field.The new method eliminates the need for separation of roots from soil and permits large-scale phenotyping of root responses to drought or other environmental and disease stresses in the field.

View Article: PubMed Central - PubMed

Affiliation: Australian Centre for Plant Functional Genomics, The University of Adelaide, Waite Campus, Glen Osmond, South Australia, 5064, Australia.

ABSTRACT
Root systems are critical for water and nutrient acquisition by crops. Current methods measuring root biomass and length are slow and labour-intensive for studying root responses to environmental stresses in the field. Here, we report the development of a method that measures changes in the root DNA concentration in soil and detects root responses to drought in controlled environment and field trials. To allow comparison of soil DNA concentrations from different wheat genotypes, we also developed a procedure for correcting genotypic differences in the copy number of the target DNA sequence. The new method eliminates the need for separation of roots from soil and permits large-scale phenotyping of root responses to drought or other environmental and disease stresses in the field.

Show MeSH
Related in: MedlinePlus