Genetics, physiology, proteomics of quality protein maize inbred lines under drought and heat stress
- Authors: Pfunde, Cleopatra Nyaradzo
- Date: 2016
- Subjects: Corn Drought-tolerant plants Proteomics
- Language: English
- Type: Thesis , Masters/Doctoral , Crop Science
- Identifier: http://hdl.handle.net/10353/11696 , vital:39098
- Description: When plants perceive heat stress alone or in combination with drought, they activate several responses to synchronize development and molecular activities to ensure survival. In this study, a multidiscipline approach was adopted to investigate the plant genetics, proteomics and physiology for understanding the mechanisms underlying the adaptive response to combined drought and heat stress. The quality protein maize (QPM) inbred lines used had varied degrees of tolerance to both heat stress alone and in combination with drought. The overall objective of this study was to identify drought and heat tolerant germplasm that combine well under drought and heat stress for use in breeding programs to develop new QPM varieties. Initially, the genetic diversity among available inbred lines was estimated using two molecular markers, simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs). The results indicated that moderate diversity exists among the QPM inbred lines which can be exploited to generate high performing hybrids. Cross combinations between QS26 and CIM2 and those between QS6 and CIM14 were selected as crosses likely to give substantial heterosis in the field because of the high genetic distances that were found between them using SNPs. Screening of inbred lines for drought and heat tolerance was done at the early germination and seedling stages. Differences in heat tolerance at both stages of growth were observed. Heat tolerant inbred lines at early germination stage had longer coleoptiles and radicles after recovery. The following inbred lines exhibited acquired thermo-tolerance, QS6, CIM7, QS17 and QS19, while CIM12, QS1, CIM21 and QS32 showed basal thermo-tolerance. At the early seedling stage inbred lines with significant differences (P< 0.001) in morpho-physiological traits were noted under heat stress alone and under combined drought and heat stress. However, no significant differences were noted for morpho-physiological measurements in the control treatment. Electron transport rate, sub-stomatal CO2, and stomatal conductance were significantly (P<0.001) reduced in response to combined drought and heat stress. Thus, drought coupled with heat stress resulted in stomatal closure, decreased photosynthetic rate, enhanced respiration and increased leaf temperature for susceptible inbred lines. The combined effects of drought and heat stress were more deleterious than the individual effect of heat stress alone as indicated by the reduction in dry weight. Based on the physiological parameters, biomass and tolerance indices, inbred lines QS22, CIM18 and QS6 showed tolerance to combined drought and heat stress, while CIM18, QS14 and QS30 exhibited thermo-tolerance. Inbred line CIM18 showed cross tolerance to combined drought and heat stress, and to heat stress alone. Further investigations into the comparative proteomic analysis of CIM18 (tolerant) with QS21 (susceptible) under combined drought and heat stress at seedling stage revealed that photosynthetic and stress related proteins were upregulated in CIM18. In addition, the abundance of heat shock proteins such as HSP70 and sHSP 17.8 kDa meant that CIM18 was better able to maintain cellular homeostasis and metabolism. The presence of metabolism related proteins meant that the energy needs of photosynthesis and other activities were met under drought and heat stress. Anti-oxidant enzymes such as glutathione s transferase were responsible for detoxification of reactive oxygen species induced by oxidative stress. These proteins mediated tolerance to combined stress in CIM18. The proteins that were micro-sequenced have previously been identified in individual drought and heat stress studies. The expression of these proteins under combined stress indicated that mechanisms from both stresses were pooled to confer tolerance in CIM18. Accordingly, the study proposes that CIM18 is a drought and heat tolerant QPM inbred line and is recommended for inclusion in breeding programs aimed at the development of drought and heat tolerant varieties. QPM has the potential to improve human nutrition in several rural households. The combining ability analysis of inbred lines revealed that GCA effects under combined drought and heat stress were significant for ears per plant, chlorophyll content, canopy temperature, anthesis silking interval, days to 50percent silking and ear length. Specific combining ability effects were significant for grain yield, days to 50percent anthesis, chlorophyll content and senescence. The study revealed that non-additive gene action was more important in determining grain yield under combined drought and heat stress. Inbred lines L2, L3, L7 and L3, L7 and L11 were good sources of genes for high grain yield under well-watered and combined drought and heat stress respectively. Cross combinations L3 x L13 (3.05 t/ha) and L5 x L9 (2.95 t/ha) were the best performing single cross hybrids for grain yield while cross combination L2 x L10 (1.33t/ha) was the least performing hybrid under combined stress conditions. Under well-watered conditions, the highest yielding crosses were between L2 x L13 (6.51t/ha) and L2 x L8 (6.46 t/ha). The inbred lines identified as sources of drought and heat tolerance and the high yielding cross combinations maybe useful for improving tolerance by generating hybrids or synthetic QPM maize varieties. The inbred lines which exhibited tolerance to both drought and heat stress were genetically diverse and were able to maintain photosynthesis and hence growth during stressful conditions. The ability of these genotypes to accumulate biomass or produce yield under stressful conditions highlights underlying mechanisms which need to be further exploited. While partial proteomic work was done in this study, a more comprehensive study looking into greater depth at the genes responsible for these mechanisms is paramount to fully utilize and/or transfer these genes to other desirable genotypes.
- Full Text:
- Date Issued: 2016
Genetics, physiology, proteomics of quality protein maize inbred lines under drought and heat stress
- Authors: Pfunde, Cleopatra Nyaradzo
- Date: 2016
- Subjects: Corn Drought-tolerant plants Proteomics
- Language: English
- Type: Thesis , Masters/Doctoral , Crop Science
- Identifier: http://hdl.handle.net/10353/11696 , vital:39098
- Description: When plants perceive heat stress alone or in combination with drought, they activate several responses to synchronize development and molecular activities to ensure survival. In this study, a multidiscipline approach was adopted to investigate the plant genetics, proteomics and physiology for understanding the mechanisms underlying the adaptive response to combined drought and heat stress. The quality protein maize (QPM) inbred lines used had varied degrees of tolerance to both heat stress alone and in combination with drought. The overall objective of this study was to identify drought and heat tolerant germplasm that combine well under drought and heat stress for use in breeding programs to develop new QPM varieties. Initially, the genetic diversity among available inbred lines was estimated using two molecular markers, simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs). The results indicated that moderate diversity exists among the QPM inbred lines which can be exploited to generate high performing hybrids. Cross combinations between QS26 and CIM2 and those between QS6 and CIM14 were selected as crosses likely to give substantial heterosis in the field because of the high genetic distances that were found between them using SNPs. Screening of inbred lines for drought and heat tolerance was done at the early germination and seedling stages. Differences in heat tolerance at both stages of growth were observed. Heat tolerant inbred lines at early germination stage had longer coleoptiles and radicles after recovery. The following inbred lines exhibited acquired thermo-tolerance, QS6, CIM7, QS17 and QS19, while CIM12, QS1, CIM21 and QS32 showed basal thermo-tolerance. At the early seedling stage inbred lines with significant differences (P< 0.001) in morpho-physiological traits were noted under heat stress alone and under combined drought and heat stress. However, no significant differences were noted for morpho-physiological measurements in the control treatment. Electron transport rate, sub-stomatal CO2, and stomatal conductance were significantly (P<0.001) reduced in response to combined drought and heat stress. Thus, drought coupled with heat stress resulted in stomatal closure, decreased photosynthetic rate, enhanced respiration and increased leaf temperature for susceptible inbred lines. The combined effects of drought and heat stress were more deleterious than the individual effect of heat stress alone as indicated by the reduction in dry weight. Based on the physiological parameters, biomass and tolerance indices, inbred lines QS22, CIM18 and QS6 showed tolerance to combined drought and heat stress, while CIM18, QS14 and QS30 exhibited thermo-tolerance. Inbred line CIM18 showed cross tolerance to combined drought and heat stress, and to heat stress alone. Further investigations into the comparative proteomic analysis of CIM18 (tolerant) with QS21 (susceptible) under combined drought and heat stress at seedling stage revealed that photosynthetic and stress related proteins were upregulated in CIM18. In addition, the abundance of heat shock proteins such as HSP70 and sHSP 17.8 kDa meant that CIM18 was better able to maintain cellular homeostasis and metabolism. The presence of metabolism related proteins meant that the energy needs of photosynthesis and other activities were met under drought and heat stress. Anti-oxidant enzymes such as glutathione s transferase were responsible for detoxification of reactive oxygen species induced by oxidative stress. These proteins mediated tolerance to combined stress in CIM18. The proteins that were micro-sequenced have previously been identified in individual drought and heat stress studies. The expression of these proteins under combined stress indicated that mechanisms from both stresses were pooled to confer tolerance in CIM18. Accordingly, the study proposes that CIM18 is a drought and heat tolerant QPM inbred line and is recommended for inclusion in breeding programs aimed at the development of drought and heat tolerant varieties. QPM has the potential to improve human nutrition in several rural households. The combining ability analysis of inbred lines revealed that GCA effects under combined drought and heat stress were significant for ears per plant, chlorophyll content, canopy temperature, anthesis silking interval, days to 50percent silking and ear length. Specific combining ability effects were significant for grain yield, days to 50percent anthesis, chlorophyll content and senescence. The study revealed that non-additive gene action was more important in determining grain yield under combined drought and heat stress. Inbred lines L2, L3, L7 and L3, L7 and L11 were good sources of genes for high grain yield under well-watered and combined drought and heat stress respectively. Cross combinations L3 x L13 (3.05 t/ha) and L5 x L9 (2.95 t/ha) were the best performing single cross hybrids for grain yield while cross combination L2 x L10 (1.33t/ha) was the least performing hybrid under combined stress conditions. Under well-watered conditions, the highest yielding crosses were between L2 x L13 (6.51t/ha) and L2 x L8 (6.46 t/ha). The inbred lines identified as sources of drought and heat tolerance and the high yielding cross combinations maybe useful for improving tolerance by generating hybrids or synthetic QPM maize varieties. The inbred lines which exhibited tolerance to both drought and heat stress were genetically diverse and were able to maintain photosynthesis and hence growth during stressful conditions. The ability of these genotypes to accumulate biomass or produce yield under stressful conditions highlights underlying mechanisms which need to be further exploited. While partial proteomic work was done in this study, a more comprehensive study looking into greater depth at the genes responsible for these mechanisms is paramount to fully utilize and/or transfer these genes to other desirable genotypes.
- Full Text:
- Date Issued: 2016
Parent characterization of quality protein maize (Zea mays L.) and combining ability for tolerance to drought stress
- Authors: Pfunde, Cleopatra Nyaradzo
- Date: 2012
- Subjects: Corn -- Quality , Corn as food , Corn -- Effect of stress on , Corn -- Effect of drought on , Cluster analysis , Crops -- Effect of drought on , Corn -- Drought tolerance , Corn -- Breeding , Crops -- Drought tolerance
- Language: English
- Type: Thesis , Masters , MSc Agric (Crop Science)
- Identifier: vital:11869 , http://hdl.handle.net/10353/d1007536 , Corn -- Quality , Corn as food , Corn -- Effect of stress on , Corn -- Effect of drought on , Cluster analysis , Crops -- Effect of drought on , Corn -- Drought tolerance , Corn -- Breeding , Crops -- Drought tolerance
- Description: Quality protein maize (QPM) has enhanced levels of two essential amino acids, lysine and tryptophan compared to normal maize. This makes QPM an important cereal crop in communities where maize is a staple crop. The main abiotic factor to QPM production is drought stress. Little information is available on the effect of drought stress on QPM. Therefore, the objectives of this study were to: (i) conduct diversity analysis of QPM inbred lines using morpho-agronomic and simple sequence repeat markers, (ii) screen available QPM inbred lines and F1 progeny for tolerance to seedling drought stress, (iii) determine the combining ability and type of gene action of QPM inbred lines for tolerance to seedling drought stress, grain yield and endosperm modification. The study was conducted in South Africa, at the University of Fort Hare. Morphological characterisation of 21 inbred lines was done using quantitative and qualitative traits. A randomised complete block design with three replicates was used for characterizing the inbred lines in the field. Genstat statistical software, version 12 (Genstat ®, 2009) was used for analysis of variance (ANOVA) and descriptive statistics. Analysis of variance was performed on all quantitative data for morphological traits. Data for qualitative traits was tabulated in their nominal classes. Traits that contributed most to the variation were days to anthesis, days to silking, anthesis-silking interval, plant height, number of kernel rows, ear length and grain yield. Cluster analysis grouped the inbred lines into three main clusters. The first cluster was characterised by tall and average yielding lines, while the second cluster showed the least anthesis-silking interval, and had the highest yield. Cluster three consisted of lines that were early maturing, but were the least yielding. Genetic distances between maize inbred lines were quantified by using 27 simple sequence repeat markers. The genetic distances between genotypes was computed using Roger’s (1972) genetic distances. Cluster analysis was then carried out using the neighbour-joining tree method using Power Marker software version 3.25. A dendrogram generated from the genetic study of the inbred lines revealed three groups that concurred with expectations based upon pedigree data. These groups were not identical to the groups generated using morpho-agronomic characterisation. Twenty one QPM inbred lines were crossed using a North Carolina design II mating scheme. These were divided into seven sets, each with three inbred lines. The three inbred lines in one set were used as females and crossed with three inbred lines in another set consisting of males. Each inbred line was used as a female in one set, and as a male in a second set. Sixty three hybrids (7 sets x 9 hybrids) were formed and evaluated in October 2011, using a 6x8 alpha-lattice incomplete block design with three replicates under glasshouse and optimum field conditions. A randomised complete block design with three replicates was used for the 21 parental inbred lines. Traits recorded for the glasshouse study were, canopy temperature, chlorophyll content, leaf roll, stem diameter, plant height, leaf number, leaf area, fresh and dry root and shoot weights. Data for the various traits for each environment, 25 percent (stress treatment) and 75 percent (non-stress) of field capacity, were subjected to analysis of variance using the unbalanced treatment design in Genstat statistical package Edition 12. Where varietal differences were found, means were separated using Tukey’s test. Genetic analyses for grain yield and agronomic traits were performed using a fixed effects model in JMP 10 following Residual Maximum Likelihood procedure (REML). From the results, inbred lines that were not previously classified into heterotic groups and drought tolerance categories were classified based on their total dry weight performance and drought susceptibility index. Inbred lines L18, L9, L8, L6 and L3, in order of their drought tolerance index were the best performers under greenhouse conditions and could be recommended for breeding new varieties that are tolerant to seedling drought stress. Evaluation of maize seedlings tolerant to drought stress under glasshouse conditions revealed that cross combination L18 x L11 was drought tolerant, while cross L20 x L7 was susceptible. Total dry weight was used as the major criteria for classifying F1 maize seedlings as being resistant or susceptible. General combining ability effects accounted for 67.43 percent of the genetic variation for total dry weight, while specific combining ability effects contributed 37.57 percent. This indicated that additive gene effects were more important than non-additive gene action in controlling this trait. In the field study (non-drought), the experimental design was a 6x8 alpha lattice incomplete block design with three replicates. On an adjacent field a randomised complete block design with three replicates was used to evaluate the parental inbred lines. The following variables were recorded: plant height, ear height, ears per plant, endosperm modification, days to silking and days to anthesis, anthesis-silking interval, number of kernels per row, number of rows per ear and grain yield. General analyses for the incomplete lattice block design and randomised complete block design for hybrid and inbred data respectively were performed using JMP 10 statistical software. Means were separated using the Tukey's test. Genetic analyses of data for grain yield and agronomic traits were conducted using a fixed effects model using REML in JMP 10. The importance of both GCA (51 percent) and SCA (49 percent) was observed for grain yield. A preponderance of GCA existed for ear height, days to anthesis, anthesis-silking interval, ears per plant and number of kernels per row, indicating that predominantly, additive gene effects controlled hybrid performance under optimum field conditions. The highest heritability was observed for days to silking (48.27 percent) suggesting that yield could be improved through selection for this trait. Under field conditions, variation in time to maturity was observed. This implies that these inbred lines can be recommended for utilisation in different agro-ecologies. Early maturing lines such as L18 can be used to introduce earliness in local cultivars, while early maturing single crosses such as L18 x L2, L5 x L9, L3 x L4 and L2 x L21 could be recommended for maize growers in drought prone areas such as the former Ciskei. Single crosses L18xL11, L16xL18, L8xL21 and L9xL6 had good tolerance to seedling drought stress. On the other hand, single crosses L18xL11 and L11xL13 had high grain yield and good endosperm modification. All these single crosses could be recommended for commercial production after evaluation across locations in the Eastern Cape Province. Alternatively they can be crossed with other superior inbreds to generate three or four way hybrids, which could then be evaluated for potential use by farmers in the Eastern Cape.
- Full Text:
- Date Issued: 2012
- Authors: Pfunde, Cleopatra Nyaradzo
- Date: 2012
- Subjects: Corn -- Quality , Corn as food , Corn -- Effect of stress on , Corn -- Effect of drought on , Cluster analysis , Crops -- Effect of drought on , Corn -- Drought tolerance , Corn -- Breeding , Crops -- Drought tolerance
- Language: English
- Type: Thesis , Masters , MSc Agric (Crop Science)
- Identifier: vital:11869 , http://hdl.handle.net/10353/d1007536 , Corn -- Quality , Corn as food , Corn -- Effect of stress on , Corn -- Effect of drought on , Cluster analysis , Crops -- Effect of drought on , Corn -- Drought tolerance , Corn -- Breeding , Crops -- Drought tolerance
- Description: Quality protein maize (QPM) has enhanced levels of two essential amino acids, lysine and tryptophan compared to normal maize. This makes QPM an important cereal crop in communities where maize is a staple crop. The main abiotic factor to QPM production is drought stress. Little information is available on the effect of drought stress on QPM. Therefore, the objectives of this study were to: (i) conduct diversity analysis of QPM inbred lines using morpho-agronomic and simple sequence repeat markers, (ii) screen available QPM inbred lines and F1 progeny for tolerance to seedling drought stress, (iii) determine the combining ability and type of gene action of QPM inbred lines for tolerance to seedling drought stress, grain yield and endosperm modification. The study was conducted in South Africa, at the University of Fort Hare. Morphological characterisation of 21 inbred lines was done using quantitative and qualitative traits. A randomised complete block design with three replicates was used for characterizing the inbred lines in the field. Genstat statistical software, version 12 (Genstat ®, 2009) was used for analysis of variance (ANOVA) and descriptive statistics. Analysis of variance was performed on all quantitative data for morphological traits. Data for qualitative traits was tabulated in their nominal classes. Traits that contributed most to the variation were days to anthesis, days to silking, anthesis-silking interval, plant height, number of kernel rows, ear length and grain yield. Cluster analysis grouped the inbred lines into three main clusters. The first cluster was characterised by tall and average yielding lines, while the second cluster showed the least anthesis-silking interval, and had the highest yield. Cluster three consisted of lines that were early maturing, but were the least yielding. Genetic distances between maize inbred lines were quantified by using 27 simple sequence repeat markers. The genetic distances between genotypes was computed using Roger’s (1972) genetic distances. Cluster analysis was then carried out using the neighbour-joining tree method using Power Marker software version 3.25. A dendrogram generated from the genetic study of the inbred lines revealed three groups that concurred with expectations based upon pedigree data. These groups were not identical to the groups generated using morpho-agronomic characterisation. Twenty one QPM inbred lines were crossed using a North Carolina design II mating scheme. These were divided into seven sets, each with three inbred lines. The three inbred lines in one set were used as females and crossed with three inbred lines in another set consisting of males. Each inbred line was used as a female in one set, and as a male in a second set. Sixty three hybrids (7 sets x 9 hybrids) were formed and evaluated in October 2011, using a 6x8 alpha-lattice incomplete block design with three replicates under glasshouse and optimum field conditions. A randomised complete block design with three replicates was used for the 21 parental inbred lines. Traits recorded for the glasshouse study were, canopy temperature, chlorophyll content, leaf roll, stem diameter, plant height, leaf number, leaf area, fresh and dry root and shoot weights. Data for the various traits for each environment, 25 percent (stress treatment) and 75 percent (non-stress) of field capacity, were subjected to analysis of variance using the unbalanced treatment design in Genstat statistical package Edition 12. Where varietal differences were found, means were separated using Tukey’s test. Genetic analyses for grain yield and agronomic traits were performed using a fixed effects model in JMP 10 following Residual Maximum Likelihood procedure (REML). From the results, inbred lines that were not previously classified into heterotic groups and drought tolerance categories were classified based on their total dry weight performance and drought susceptibility index. Inbred lines L18, L9, L8, L6 and L3, in order of their drought tolerance index were the best performers under greenhouse conditions and could be recommended for breeding new varieties that are tolerant to seedling drought stress. Evaluation of maize seedlings tolerant to drought stress under glasshouse conditions revealed that cross combination L18 x L11 was drought tolerant, while cross L20 x L7 was susceptible. Total dry weight was used as the major criteria for classifying F1 maize seedlings as being resistant or susceptible. General combining ability effects accounted for 67.43 percent of the genetic variation for total dry weight, while specific combining ability effects contributed 37.57 percent. This indicated that additive gene effects were more important than non-additive gene action in controlling this trait. In the field study (non-drought), the experimental design was a 6x8 alpha lattice incomplete block design with three replicates. On an adjacent field a randomised complete block design with three replicates was used to evaluate the parental inbred lines. The following variables were recorded: plant height, ear height, ears per plant, endosperm modification, days to silking and days to anthesis, anthesis-silking interval, number of kernels per row, number of rows per ear and grain yield. General analyses for the incomplete lattice block design and randomised complete block design for hybrid and inbred data respectively were performed using JMP 10 statistical software. Means were separated using the Tukey's test. Genetic analyses of data for grain yield and agronomic traits were conducted using a fixed effects model using REML in JMP 10. The importance of both GCA (51 percent) and SCA (49 percent) was observed for grain yield. A preponderance of GCA existed for ear height, days to anthesis, anthesis-silking interval, ears per plant and number of kernels per row, indicating that predominantly, additive gene effects controlled hybrid performance under optimum field conditions. The highest heritability was observed for days to silking (48.27 percent) suggesting that yield could be improved through selection for this trait. Under field conditions, variation in time to maturity was observed. This implies that these inbred lines can be recommended for utilisation in different agro-ecologies. Early maturing lines such as L18 can be used to introduce earliness in local cultivars, while early maturing single crosses such as L18 x L2, L5 x L9, L3 x L4 and L2 x L21 could be recommended for maize growers in drought prone areas such as the former Ciskei. Single crosses L18xL11, L16xL18, L8xL21 and L9xL6 had good tolerance to seedling drought stress. On the other hand, single crosses L18xL11 and L11xL13 had high grain yield and good endosperm modification. All these single crosses could be recommended for commercial production after evaluation across locations in the Eastern Cape Province. Alternatively they can be crossed with other superior inbreds to generate three or four way hybrids, which could then be evaluated for potential use by farmers in the Eastern Cape.
- Full Text:
- Date Issued: 2012
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