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Fine mapping of the Bsr1 barley stripe mosaic virus resistance gene in the model grass Brachypodium distachyon.

Cui Y, Lee MY, Huo N, Bragg J, Yan L, Yuan C, Li C, Holditch SJ, Xie J, Luo MC, Li D, Yu J, Martin J, Schackwitz W, Gu YQ, Vogel JP, Jackson AO, Liu Z, Garvin DF - PLoS ONE (2012)

Bottom Line: We generated an F(6:7) recombinant inbred line (RIL) population from a cross between Bd3-1 and Bd21 and used the RILs, and an F(2) population of a second Bd21 × Bd3-1 cross to evaluate the inheritance of resistance.Our study demonstrates the power of using RILs to rapidly map the genetic determinants of BSMV resistance in Brachypodium.Moreover, the RILs and their associated genetic map, when combined with the complete genomic sequence of Brachypodium, provide new resources for genetic analyses of many other traits.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Agro-Biotechnology, China Agricultural University, Beijing, China.

ABSTRACT
The ND18 strain of Barley stripe mosaic virus (BSMV) infects several lines of Brachypodium distachyon, a recently developed model system for genomics research in cereals. Among the inbred lines tested, Bd3-1 is highly resistant at 20 to 25 °C, whereas Bd21 is susceptible and infection results in an intense mosaic phenotype accompanied by high levels of replicating virus. We generated an F(6:7) recombinant inbred line (RIL) population from a cross between Bd3-1 and Bd21 and used the RILs, and an F(2) population of a second Bd21 × Bd3-1 cross to evaluate the inheritance of resistance. The results indicate that resistance segregates as expected for a single dominant gene, which we have designated Barley stripe mosaic virus resistance 1 (Bsr1). We constructed a genetic linkage map of the RIL population using SNP markers to map this gene to within 705 Kb of the distal end of the top of chromosome 3. Additional CAPS and Indel markers were used to fine map Bsr1 to a 23 Kb interval containing five putative genes. Our study demonstrates the power of using RILs to rapidly map the genetic determinants of BSMV resistance in Brachypodium. Moreover, the RILs and their associated genetic map, when combined with the complete genomic sequence of Brachypodium, provide new resources for genetic analyses of many other traits.

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Disease responses of Brachypodium lines Bd3-1, Bd21 and Bd21-3 to infection with BSMV ND18.(A–C) Uninfected Bd3-1 and inoculated Bd-3-1, Bd21 and Bd21-3 at 12 dpi. (A) Uninfected Bd3-1 plants remained green and continued to grow rapidly, as was typical of uninfected Bd21 and Bd21-3 plants. Nd18 inoculated Bd3-1 plants failed to develop symptoms and had the same general appearance as their uninoculated counterparts. In contrast, Bd21 and Bd21-3 inoculated plants developed visible mosaic symptoms on emerging leaves by 7 days post inoculation (dpi) and the symptoms remain visible until at least 20 dpi. (B) Western blots to determine the presence of the 22 KD BSMV coat protein in leaf extracts from the first emerging leaf of uninoculated and inoculated plants at 6 dpi. (C) RT-PCR analyses of leaf extracts taken at 21 dpi from the lines shown in the top panel. A forward primer complementary to the 3′ end of BSMV RNAs and a reverse primer of the same polarity as the γb gene were designed to produce an ∼800 nt product. (D–E) Chronic disease symptoms on Brachypodium lines inoculated with BSMV ND18. (D) Bd3-1 and Bd21 at 25 dpi. Note stunting of Bd21 compared to Bd3-1. (E) Healthy Bd3-1 and Bd 3-1 and Bd21 at 55 dpi. Note: Uninoculated plants and inoculated Bd3-1 plants have a similar growth characteristics and seed population, but Bd21 plants are stunted and fail to flower or set seeds.
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pone-0038333-g002: Disease responses of Brachypodium lines Bd3-1, Bd21 and Bd21-3 to infection with BSMV ND18.(A–C) Uninfected Bd3-1 and inoculated Bd-3-1, Bd21 and Bd21-3 at 12 dpi. (A) Uninfected Bd3-1 plants remained green and continued to grow rapidly, as was typical of uninfected Bd21 and Bd21-3 plants. Nd18 inoculated Bd3-1 plants failed to develop symptoms and had the same general appearance as their uninoculated counterparts. In contrast, Bd21 and Bd21-3 inoculated plants developed visible mosaic symptoms on emerging leaves by 7 days post inoculation (dpi) and the symptoms remain visible until at least 20 dpi. (B) Western blots to determine the presence of the 22 KD BSMV coat protein in leaf extracts from the first emerging leaf of uninoculated and inoculated plants at 6 dpi. (C) RT-PCR analyses of leaf extracts taken at 21 dpi from the lines shown in the top panel. A forward primer complementary to the 3′ end of BSMV RNAs and a reverse primer of the same polarity as the γb gene were designed to produce an ∼800 nt product. (D–E) Chronic disease symptoms on Brachypodium lines inoculated with BSMV ND18. (D) Bd3-1 and Bd21 at 25 dpi. Note stunting of Bd21 compared to Bd3-1. (E) Healthy Bd3-1 and Bd 3-1 and Bd21 at 55 dpi. Note: Uninoculated plants and inoculated Bd3-1 plants have a similar growth characteristics and seed population, but Bd21 plants are stunted and fail to flower or set seeds.

Mentions: Brachypodium inbred lines Bd3-1, Bd21 and Bd21-3, all from Iraq, are of particular interest because of the resources available for these lines. The phenotypic responses of Bd3-1, Bd21 and Bd21-3 on emerging leaves inoculated with ND18 along with western blot analyses of coat protein, and RT-PCR of sequences common to the 3′ sequences of the three genomic RNAs of the virus are shown in Figure 2A–C. These experiments revealed that Bd21 and Bd21-3 are susceptible to ND18 because both inbred lines developed mosaic symptoms within 7 dpi and accumulated high levels of virus RNA and protein, whereas Bd3-1 failed to develop symptoms and did not contain detectable viral RNA or protein. After 3 weeks, substantial stunting of infected plants was evident and the plants failed to set seeds upon maturity (Figure 2D and 2E). In contrast, Bd3-1 failed to develop mosaic symptoms and its growth and seed production was indistinguishable from uninoculated plants.


Fine mapping of the Bsr1 barley stripe mosaic virus resistance gene in the model grass Brachypodium distachyon.

Cui Y, Lee MY, Huo N, Bragg J, Yan L, Yuan C, Li C, Holditch SJ, Xie J, Luo MC, Li D, Yu J, Martin J, Schackwitz W, Gu YQ, Vogel JP, Jackson AO, Liu Z, Garvin DF - PLoS ONE (2012)

Disease responses of Brachypodium lines Bd3-1, Bd21 and Bd21-3 to infection with BSMV ND18.(A–C) Uninfected Bd3-1 and inoculated Bd-3-1, Bd21 and Bd21-3 at 12 dpi. (A) Uninfected Bd3-1 plants remained green and continued to grow rapidly, as was typical of uninfected Bd21 and Bd21-3 plants. Nd18 inoculated Bd3-1 plants failed to develop symptoms and had the same general appearance as their uninoculated counterparts. In contrast, Bd21 and Bd21-3 inoculated plants developed visible mosaic symptoms on emerging leaves by 7 days post inoculation (dpi) and the symptoms remain visible until at least 20 dpi. (B) Western blots to determine the presence of the 22 KD BSMV coat protein in leaf extracts from the first emerging leaf of uninoculated and inoculated plants at 6 dpi. (C) RT-PCR analyses of leaf extracts taken at 21 dpi from the lines shown in the top panel. A forward primer complementary to the 3′ end of BSMV RNAs and a reverse primer of the same polarity as the γb gene were designed to produce an ∼800 nt product. (D–E) Chronic disease symptoms on Brachypodium lines inoculated with BSMV ND18. (D) Bd3-1 and Bd21 at 25 dpi. Note stunting of Bd21 compared to Bd3-1. (E) Healthy Bd3-1 and Bd 3-1 and Bd21 at 55 dpi. Note: Uninoculated plants and inoculated Bd3-1 plants have a similar growth characteristics and seed population, but Bd21 plants are stunted and fail to flower or set seeds.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3366947&req=5

pone-0038333-g002: Disease responses of Brachypodium lines Bd3-1, Bd21 and Bd21-3 to infection with BSMV ND18.(A–C) Uninfected Bd3-1 and inoculated Bd-3-1, Bd21 and Bd21-3 at 12 dpi. (A) Uninfected Bd3-1 plants remained green and continued to grow rapidly, as was typical of uninfected Bd21 and Bd21-3 plants. Nd18 inoculated Bd3-1 plants failed to develop symptoms and had the same general appearance as their uninoculated counterparts. In contrast, Bd21 and Bd21-3 inoculated plants developed visible mosaic symptoms on emerging leaves by 7 days post inoculation (dpi) and the symptoms remain visible until at least 20 dpi. (B) Western blots to determine the presence of the 22 KD BSMV coat protein in leaf extracts from the first emerging leaf of uninoculated and inoculated plants at 6 dpi. (C) RT-PCR analyses of leaf extracts taken at 21 dpi from the lines shown in the top panel. A forward primer complementary to the 3′ end of BSMV RNAs and a reverse primer of the same polarity as the γb gene were designed to produce an ∼800 nt product. (D–E) Chronic disease symptoms on Brachypodium lines inoculated with BSMV ND18. (D) Bd3-1 and Bd21 at 25 dpi. Note stunting of Bd21 compared to Bd3-1. (E) Healthy Bd3-1 and Bd 3-1 and Bd21 at 55 dpi. Note: Uninoculated plants and inoculated Bd3-1 plants have a similar growth characteristics and seed population, but Bd21 plants are stunted and fail to flower or set seeds.
Mentions: Brachypodium inbred lines Bd3-1, Bd21 and Bd21-3, all from Iraq, are of particular interest because of the resources available for these lines. The phenotypic responses of Bd3-1, Bd21 and Bd21-3 on emerging leaves inoculated with ND18 along with western blot analyses of coat protein, and RT-PCR of sequences common to the 3′ sequences of the three genomic RNAs of the virus are shown in Figure 2A–C. These experiments revealed that Bd21 and Bd21-3 are susceptible to ND18 because both inbred lines developed mosaic symptoms within 7 dpi and accumulated high levels of virus RNA and protein, whereas Bd3-1 failed to develop symptoms and did not contain detectable viral RNA or protein. After 3 weeks, substantial stunting of infected plants was evident and the plants failed to set seeds upon maturity (Figure 2D and 2E). In contrast, Bd3-1 failed to develop mosaic symptoms and its growth and seed production was indistinguishable from uninoculated plants.

Bottom Line: We generated an F(6:7) recombinant inbred line (RIL) population from a cross between Bd3-1 and Bd21 and used the RILs, and an F(2) population of a second Bd21 × Bd3-1 cross to evaluate the inheritance of resistance.Our study demonstrates the power of using RILs to rapidly map the genetic determinants of BSMV resistance in Brachypodium.Moreover, the RILs and their associated genetic map, when combined with the complete genomic sequence of Brachypodium, provide new resources for genetic analyses of many other traits.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Agro-Biotechnology, China Agricultural University, Beijing, China.

ABSTRACT
The ND18 strain of Barley stripe mosaic virus (BSMV) infects several lines of Brachypodium distachyon, a recently developed model system for genomics research in cereals. Among the inbred lines tested, Bd3-1 is highly resistant at 20 to 25 °C, whereas Bd21 is susceptible and infection results in an intense mosaic phenotype accompanied by high levels of replicating virus. We generated an F(6:7) recombinant inbred line (RIL) population from a cross between Bd3-1 and Bd21 and used the RILs, and an F(2) population of a second Bd21 × Bd3-1 cross to evaluate the inheritance of resistance. The results indicate that resistance segregates as expected for a single dominant gene, which we have designated Barley stripe mosaic virus resistance 1 (Bsr1). We constructed a genetic linkage map of the RIL population using SNP markers to map this gene to within 705 Kb of the distal end of the top of chromosome 3. Additional CAPS and Indel markers were used to fine map Bsr1 to a 23 Kb interval containing five putative genes. Our study demonstrates the power of using RILs to rapidly map the genetic determinants of BSMV resistance in Brachypodium. Moreover, the RILs and their associated genetic map, when combined with the complete genomic sequence of Brachypodium, provide new resources for genetic analyses of many other traits.

Show MeSH
Related in: MedlinePlus