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Expression Profiling Coupled with In-silico Mapping Identifies Candidate Genes for Reducing Aflatoxin Accumulation in Maize

View Article: PubMed Central - PubMed

ABSTRACT

Aflatoxin, produced by Aspergillus flavus, is hazardous to health of humans and livestock. The lack of information about large effect QTL for resistance to aflatoxin accumulation is a major obstacle to employ marker-assisted selection for maize improvement. The understanding of resistance mechanisms of the host plant and the associated genes is necessary for improving resistance to A. flavus infection. A suppression subtraction hybridization (SSH) cDNA library was made using the developing kernels of Mp715 (resistant inbred) and B73 (susceptible inbred) and 480 randomly selected cDNA clones were sequenced to identify differentially expressed genes (DEGs) in response to A. flavus infection and map these clones onto the corn genome by in-silico mapping. A total of 267 unigenes were identified and majority of genes were related to metabolism, stress response, and disease resistance. Based on the reverse northern hybridization experiment, 26 DEGs were selected for semi-quantitative RT-PCR analysis in seven inbreds with variable resistance to aflatoxin accumulation at two time points after A. flavus inoculation. Most of these genes were highly expressed in resistant inbreds. Quantitative RT-PCR analysis validated upregulation of PR-4, DEAD-box RNA helicase, and leucine rich repeat family protein in resistant inbreds. Fifty-six unigenes, which were placed on linkage map through in-silico mapping, overlapped the QTL regions for resistance to aflatoxin accumulation identified in a mapping population derived from the cross between B73 and Mp715. Since majority of these mapped genes were related to disease resistance, stress response, and metabolism, these should be ideal candidates to investigate host pathogen interaction and to reduce aflatoxin accumulation in maize.

No MeSH data available.


Co-localization of differentially expressed genes from the SSH library with the QTL for resistance to aflatoxin accumulation in an F2:3 population derived from B73 × Mp715 (Dhakal et al., 2016). Red, blue, and black bars represent QTL for aflatoxin resistance during 2012, 2013, and combined analysis, respectively. Clone ID in red font were used in RT-PCR for expression studies on various inbred lines. Green bars represent the location of QTL identified by Brooks et al. (2005). Detail information about the genes is provided in Table S2.
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Figure 5: Co-localization of differentially expressed genes from the SSH library with the QTL for resistance to aflatoxin accumulation in an F2:3 population derived from B73 × Mp715 (Dhakal et al., 2016). Red, blue, and black bars represent QTL for aflatoxin resistance during 2012, 2013, and combined analysis, respectively. Clone ID in red font were used in RT-PCR for expression studies on various inbred lines. Green bars represent the location of QTL identified by Brooks et al. (2005). Detail information about the genes is provided in Table S2.

Mentions: The ESTs from the SSH library were mapped on to the maize linkage map constructed from the cross B73 × Mp715 (Dhakal et al., 2016; Figure 5; Table S2). Altogether 56 genes with various physiological functions co-localized with the QTL regions. These genes showed high similarity (>95%) to the annotated genes on the reference maize genome and were distributed on different chromosomes, except chromosomes 6, 7, and 8. Majority of these genes were mapped on/near QTL for resistance to aflatoxin accumulation on chromosomes 2, 4, 5, and 10. These genes were largely related to metabolism, stress response, and disease resistance suggesting the usefulness of this integrated approach to identify candidate genes to prevent A. flavus infection.


Expression Profiling Coupled with In-silico Mapping Identifies Candidate Genes for Reducing Aflatoxin Accumulation in Maize
Co-localization of differentially expressed genes from the SSH library with the QTL for resistance to aflatoxin accumulation in an F2:3 population derived from B73 × Mp715 (Dhakal et al., 2016). Red, blue, and black bars represent QTL for aflatoxin resistance during 2012, 2013, and combined analysis, respectively. Clone ID in red font were used in RT-PCR for expression studies on various inbred lines. Green bars represent the location of QTL identified by Brooks et al. (2005). Detail information about the genes is provided in Table S2.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5382453&req=5

Figure 5: Co-localization of differentially expressed genes from the SSH library with the QTL for resistance to aflatoxin accumulation in an F2:3 population derived from B73 × Mp715 (Dhakal et al., 2016). Red, blue, and black bars represent QTL for aflatoxin resistance during 2012, 2013, and combined analysis, respectively. Clone ID in red font were used in RT-PCR for expression studies on various inbred lines. Green bars represent the location of QTL identified by Brooks et al. (2005). Detail information about the genes is provided in Table S2.
Mentions: The ESTs from the SSH library were mapped on to the maize linkage map constructed from the cross B73 × Mp715 (Dhakal et al., 2016; Figure 5; Table S2). Altogether 56 genes with various physiological functions co-localized with the QTL regions. These genes showed high similarity (>95%) to the annotated genes on the reference maize genome and were distributed on different chromosomes, except chromosomes 6, 7, and 8. Majority of these genes were mapped on/near QTL for resistance to aflatoxin accumulation on chromosomes 2, 4, 5, and 10. These genes were largely related to metabolism, stress response, and disease resistance suggesting the usefulness of this integrated approach to identify candidate genes to prevent A. flavus infection.

View Article: PubMed Central - PubMed

ABSTRACT

Aflatoxin, produced by Aspergillus flavus, is hazardous to health of humans and livestock. The lack of information about large effect QTL for resistance to aflatoxin accumulation is a major obstacle to employ marker-assisted selection for maize improvement. The understanding of resistance mechanisms of the host plant and the associated genes is necessary for improving resistance to A. flavus infection. A suppression subtraction hybridization (SSH) cDNA library was made using the developing kernels of Mp715 (resistant inbred) and B73 (susceptible inbred) and 480 randomly selected cDNA clones were sequenced to identify differentially expressed genes (DEGs) in response to A. flavus infection and map these clones onto the corn genome by in-silico mapping. A total of 267 unigenes were identified and majority of genes were related to metabolism, stress response, and disease resistance. Based on the reverse northern hybridization experiment, 26 DEGs were selected for semi-quantitative RT-PCR analysis in seven inbreds with variable resistance to aflatoxin accumulation at two time points after A. flavus inoculation. Most of these genes were highly expressed in resistant inbreds. Quantitative RT-PCR analysis validated upregulation of PR-4, DEAD-box RNA helicase, and leucine rich repeat family protein in resistant inbreds. Fifty-six unigenes, which were placed on linkage map through in-silico mapping, overlapped the QTL regions for resistance to aflatoxin accumulation identified in a mapping population derived from the cross between B73 and Mp715. Since majority of these mapped genes were related to disease resistance, stress response, and metabolism, these should be ideal candidates to investigate host pathogen interaction and to reduce aflatoxin accumulation in maize.

No MeSH data available.