CHAPTER I: INTRODUCTION
5. Combining ability and heterosis for sorghum grain yield and secondary traits across
5.2. Materials and Methods
5.2.1. Plant materials
The sorghum genotypes used in this study were obtained from the Sorghum National Breeding programme and some were originally from the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT). The two standard checks used during evaluation were varieties widely grown in the country with good grain yield performance and stability. The hybrids which produced a full set of seed after crossing were used for the evaluation (Figure 5-1). This procedure is used for hybrid development as well as in identifying male sterile seed parents (House, 1985). The parental lines were divided into two groups: cytoplasmic male sterile lines (CMS) designated as females and cytoplasmic male fertile lines (CMF) as males (Table 5-1). The CMS lines facilitated the hybrid development in sorghum improvement (Reddy et al., 2008a). Nine female lines (lines) and four male lines (testers) were used in a Line x Tester mating scheme. All crosses were successful and the resultant 36 hybrids together with two checks were evaluated in yield trials across six test environments (combination of year- location) during 2015/16 and 2016/17 seasons (Table 5-2). The 13 parents were also evaluated in four of the six test environments.
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Figure 5-1 Sorghum hybridization in a greenhouse (left) and sorghum head of one hybrid during field evaluation (right) at Sussundenga Research Station.
Table 5-1 Parents used in the line by tester crossing block for hybrid development
Parent Group Designation Group Origin
LARSVYT 46A CMS Female Line NAR/ICRISAT
8601A CMS Female Line NAR/ICRISAT
SPL 38A CMS Female Line NAR/ICRISAT
ICSA 19A CMS Female Line NAR/ICRISAT
TX 628A CMS Female Line NAR/ICRISAT
ICSA 21A CMS Female Line NAR/ICRISAT
ICSA 12A CMS Female Line NAR/ICRISAT
CK 60A CMS Female Line NAR/ICRISAT
TX 631A CMS Female Line NAR/ICRISAT
IS 14257R CMF Male Tester NAR/ICRISAT
SDS 6013R CMF Male Tester NAR/ICRISAT
MZ 2R CMF Male Tester NAR
MZ 37R CMF Male Tester NAR
NAR = National Research Sorghum Programme; cytoplasmic male sterile lines (CMS) and cytoplasmic male fertile lines (CMF)
100 5.2.2. Field evaluation sites
The trials were conducted at Chókwè and Maniquenique in the southern region, Sussundenga in the central region and Mapupulo in the northern region. These sites cover from low to mid- altitude environments (Table 5-2).
Table 5-2 Locations used for evaluation of entries over two seasons and rainfall received during the evaluation period
Location Season Code
Latitude (oS)
Longitude
(oE) Altitude (m)
Rainfall* (mm) Sussundenga** 2015/16 Sus16 19o18’ 33o15’ 635 522
Chókwè** 2015/16 Chk16 24o52’ 33o00’ 33 380
Sussundenga** 2016/17 Sus17 19o18’ 33o15’ 635 989
Chókwè** 2016/17 Chk17 24o52’ 33o00’ 33 650
Maniquenique 2016/17 Man17 24o73’ 33o53’ 13 468
Mapupulo 2016/17 Map17 13o19’ 38o86’ 534 1050
*Rainfall refers to the amount received during the crop growing season, **Parents evaluation sites
5.2.3. Experimental design and field management
The 36 experimental hybrids, along with two check hybrids were laid out in a 19 x 2 alpha lattice design with three replications in each environment. The 13 parents were also evaluated in the same field adjacent to the hybrids and were laid out in a randomized complete block design with three replications in four of the six environments. Individual plots consisted of two rows which were 5 m long, spaced 0.75 m apart and the distance between plants in a row was 0.25 m. Fertilizer was applied at recommended rates of 250 kg ha-1 NPK (12-24-12) basal fertilizer, and 150 kg ha-1 Urea (46% N) as a top-dressing fertilizer. Other cultural practices such as ploughing, disking, hand planting, hand weeding and herbicides and pesticide application were carried out at each site.
Ten plants were randomly selected from each hybrid in each replication to measure the characters such as days to 50% flowering, plant height, panicle length, number of tillering plants, number of panicles and panicle aspect. Grain yield, biomass and disease scoring were done at on per plot basis. Adjusted harvest grain yield was determined following the standard practices used by CIMMYT (CIMMYT, 1985) as presented:
Grain yield (t.ha-1) = [Grain weight (kg/plot) x 10 x (100-MOI*)/ (100-12.5) / (Plot Area)]
*MOI = Grain Moisture Content
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Due to high amounts of rainfall received in the 2016/17 season, three diseases severely affected some entries in four experimental sites. The diseases were Cercospora spp (similar to grey leaf spot (GLS) in maize), Puccinia purpurea (rust) and Colletotrichum graminicola (Cesati) Wilson (anthracnose). The disease severity was recorded based on visual assessment of the degree of damaged leaf area on scale of 1 to 5, where 1 = no disease, 2 = 1 to 5% leaf area damaged, 3 = 6 to 20% leaf area damage, 4 = 20 to 40% leaf area damaged, and 5 = severe disease with more than 40% leaf area damaged. The scores were further classified according to disease reaction type, where less than 5% leaf damage = resistant (R), 6 to 10% leaf damage = moderate resistant (MR), 10 to 20% leaf damage = moderately susceptible (MS) and more than 20 % leaf damage = susceptible (S).
5.2.4. Data analysis
The individual and combined season data were analysed using PROC GLM procedure in SAS 9.3 (SAS, 2011). Analyses of variance (ANOVA) were done first by environment with genotypes as the main effect, then a combined analysis across environments was conducted to evaluate the effect of environment, genotypes and their interactions. The calculation of combining ability effect was done only for the 36 hybrids tested. The data analysis followed a fixed effects model:
𝑌𝑖𝑗𝑘𝑙 = 𝜇 + 𝑙𝑖 + 𝑟𝑗(𝑙𝑖) + 𝑏𝑖𝑗 + 𝑚𝑘 + 𝑓𝑖 + (𝑚 ∗ 𝑓)𝑘𝑙 + (𝑙 ∗ 𝑚)𝑖𝑘 + (𝑙 ∗ 𝑓)𝑖𝑙 + (𝑙 ∗ 𝑚 ∗ 𝑓)𝑖𝑘𝑙 + 𝜀𝑖𝑗𝑘𝑙 Where: Yijkl = observed hybrid response; µ = overall mean of the population; li =effects of the ith environment; rj (li) = effects of the jth replication in the ith environment; bij = effects of the blocks in the jth replication in the ith environment; mk = effects of the kth male parent; fl = effects of the lth female parent; m*fkl = effects of the interaction between kth male and lth female parents; li*mk = interaction effects of the ith environment in kth male parent; li*fl = interaction effects of the ith environment in lth female parent; li*m*fkl = interaction effects of the ith environment and the interaction between the kth male and lth female parents; Ԑijkl = experimental error.
Estimation of general combining ability (GCA) and specific combining ability (SCA) effects were done according to Dabholkar (1999):
𝑘𝑗(𝑡𝑒𝑠𝑡𝑒𝑟) =𝑌.𝑗.
𝑟𝑙 − 𝑌…
𝑟𝑙𝑡 ; 𝑓𝑖 (𝑙𝑖𝑛𝑒) = 𝑌𝑖..
𝑟𝑡 −𝑌…
𝑟𝑙𝑡 and
102 𝑆𝑖𝑗 = 𝑌𝑖𝑗
𝑟 −𝑌𝑖. . 𝑟𝑡 −𝑌. 𝑗.
𝑟𝑙 +𝑌 … 𝑟𝑙𝑡
where k(tester) and f(line) represent the estimates of GCA effects ith lines and jth tester, respectively. Sij represents the SCA effects of i x jth cross. Yi.., Y.j. and Y… represent the sum of lines, testers and grand total of the grain yield respectively.
The “t” test was used to estimate the significance of the GCA and SCA, and standard errors were calculated as follows:
𝑆𝐸𝑘𝑗 = (𝑀𝑒
𝑟𝑙)12, 𝑗 = 1, … , 𝑗
𝑆𝐸𝑓𝑖 = (𝑀𝑒
𝑟𝑡)1/2, 𝑖 = 1, … , 𝑖
𝑆𝐸𝑖𝑗 = (𝑀𝑒 𝑟 )1/2
Where SEs are the standard errors for testers (SEkj), lines (SEfi) and crosses (SEij). Me - is the mean square of the error. Thus, t test:
𝑡𝑔 = 𝑔 − 0 𝑆𝐸𝑔
𝑡𝑠 = 𝑠𝑖𝑗 − 0 𝑆𝐸𝑖𝑗
Where tg – test significance for GCA effects and ts – test significance for SCA effects. The t test was considered significant at probability of 5% if the value was greater than 1.96 and significant at 1% if it was greater than 2.58.
Heterosis (H) analysis was performed to compare the hybrids with their parents and identify hybrid performance that exceeds the average parental performance (House, 1985). The heterosis was calculated using the following formulas:
1) Heterosis over the mid-parent:
(MPH) = [(F1 – MP)/*MP]*100, where F1 = hybrid mean performance, MP = average predicted performance for the two parents.
2) Standard heterosis:
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(HBP) = [(F1 – MT)/*MT]*100, where F1 = hybrid mean performance, MT = best check mean, checks mean, best parents mean or trial mean