Legume Research

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Legume Research, volume 45 issue 9 (september 2022) : 1074-1081

Induction of Genetic Variability in Cowpea Variety Videza for Extra Earliness and High Seed Yield using Gamma Irradiation Mutagenesis

Innocent Kwaku Dorvlo1, Stanley Akwesi Acquah2, Jonathan Okai Armah2, Jacob Teye Kutufam2, Emmanuel Afutu3, Alfred Anthony Darkwa3, Godwin Amenorpe1,2, Harry Mensah Amoatey1, Samuel Amiteye1,2
1Nuclear Agriculture and Radiation Processing, School of Nuclear and Allied Sciences, University of Ghana, Accra, Ghana.
2Biotechnology and Nuclear Agricultural Research Institute, Ghana Atomic Energy Commission, Accra, Ghana.
3Department of Crop Science, School of Agriculture, University of Cape Coast, Ghana.
  • Submitted04-03-2022|

  • Accepted30-05-2022|

  • First Online 21-06-2022|

  • doi 10.18805/LRF-685

Cite article:- Dorvlo Kwaku Innocent, Acquah Akwesi Stanley, Armah Okai Jonathan, Kutufam Teye Jacob, Afutu Emmanuel, Darkwa Anthony Alfred, Amenorpe Godwin, Amoatey Mensah Harry, Amiteye Samuel (2022). Induction of Genetic Variability in Cowpea Variety Videza for Extra Earliness and High Seed Yield using Gamma Irradiation Mutagenesis . Legume Research. 45(9): 1074-1081. doi: 10.18805/LRF-685.
Background: Cowpea yields are very low in the West African region due to the prevalence of drought periods. This work, therefore, aimed at developing cowpea genotypes that combine early maturity with high seed yield, through gamma irradiation mutagenesis. 

Methods: A farmer preferred cowpea variety Videza was improved via gamma mutagenesis. A determined lethal dose 50% (LD50) of 240 Gy was used to irradiate 2000 seeds of Videza by applying a Cobalt 60 source. Selected M1 generation plants exhibiting early maturity and high seed yield were advanced to M2 and further to M3 using Videza as parental control.

Result: Compared to the control Videza, the number of days to 90% maturity significantly decreased in putative mutants in the M2 (from 71 days in the control to between 50 and 66 days in mutants) and further decreased in the M3 where mutants matured 10-22 days earlier than the control. Significant increment in 100-seed weight per plant occurred in the M3 mutants. In the M3, 100-seed weight increased from 15.28 g in Videza to 21.71 g, the highest in mutants. Twelve putative mutants were identified that combine early maturity with higher seed yield than the control.
Cowpea production, processing and marketing are mainly undertaken by smallholder farmers (Odendo et al., 2011). Across Sub Saharan Africa, cowpea production is limited by frequent droughts that affect, especially the flowering and pod filling stages. According to Agbicodo et al., (2009), cowpea crop exposed to moisture stress at early flowering and pod filling stage results in yield losses of up to 80%. The cultivation of early maturing varieties is usually planned such that the crop could escape the drought  periods and pests infestations (Selvan et al., 2021; Ehlers and Hall, 1997).
Cowpea genetic variability is quite narrow due to the self-pollinating nature (Asare et al., 2010). Besides, in Ghana, early maturing widely grown varieties such as Sanzi and Asuntem developed through conventional breeding, lack the following consumer preferred traits: cream seed coat colour, large grain size and early maturity as well as pests and disease resistance (Owusu et al., 2018).
To circumvent the effects of severe droughts, farmers prefer early maturing crops (Fatokun et al., 2012). In most self-pollinated plants where genetic variability is inadequate, induced mutation using irradiation has been an effective tool for creating significant genetic variability. Mutagenesis is faster and more effective compared to conventional breeding (Subba et al., 2022). Induced mutation has been successfully applied to create variability and improve many useful traits in cowpea and other crops (Ahloowalia et al., 2004). The aim of this study was to develop early maturing and high seed yielding cowpea varieties using gamma irradiation.
Phenotypes created by induced mutation are generally not readily observable in the M1 generation due to resulting heterogeneous or chimeric plants (Jankowicz-Cieslak et al., 2017). It is, therefore, useful to advance M1 plants to the M2 generation and subsequent generations.  Jankowicz-Cieslak et al., (2017) recommended the start of phenotypic screening for observable traits from the M2.
Plant materials and irradiation dose used
A farmer-preferred cowpea variety, Videza was used. Videza, has a semi-erect growth habit, ovoid seed shape and smooth seed coat texture. The variety matures in 68-77 days and produces a maximum seed yield of 2261.3 kg/ha in the major rainy season (April to July) and 1375.0 kg/ha during the minor season (September to November) (Agyeman et al., 2015). The research was conducted at the Biotechnology and Nuclear Agriculture Research Institute (BNARI), Ghana, between March 2018 and April 2018. Videza was first planted at the BNARI and harvested seeds were multiplied for two consecutive generations to develop a single seed descend population of approximately 8000 seeds for irradiation. A radio-sensitivity test was carried out following established protocols (Mba et al., 2010) to determine a dose rate of 240 Gy for mass irradiation. Thirty seeds per dose were gamma irradiated in March 2019 using a Cobalt 60 source.
Treatment of seeds and raising M1, M2 and Mpopulations
About 2000 seeds were acutely irradiated at 240 Gy. Non-irradiated materials served as control. The M1 seeds were planted under normal growing conditions. The M1 field comprised 10 rows of irradiated seeds separated by a spacing of 1.50 m from three rows of control fields. Induced M1 plants with early maturity and large seed size were selected and the M2 seeds harvested individually. All M2 seeds were used to establish the M2 mutant population as described for the M1 population. Data on the M2 generation were recorded from ten plants in each progeny row. At maturity, tagged induced Mlines from each progeny line with early maturity and high seed yield were selected and M3 seeds harvested individually. The Mseeds were sown as single-plant progenies to establish the M3 generation. Induced M3 plants from each progeny line with early maturing and high seed size were harvested separately.
Number of days (ND) to 50% flowering at M2 and M3 generations
ND to 50% flowering at M2 and M3 are presented in Fig 1. In the M2, ND to 50% flowering ranged from 32 days to 46 days among the M2 lines. The earliest to attain 50% flowering was in putative mutant P3N01 while the latest was in P10N29. The control plant flowered at 47 DAP. In the M3, ND to 50% flowering ranged from 30 days to 36 days among the M3 lines. Mutants P1N02#4, P3N01#5, P4N03#2 and P4N14#7 flowered on day 30 after planting. The latest to attain 50% flowering were P2N09#16 and P6N10#19. The control plants in the M3 attained 50% flowering at 46 DAP.

Fig 1: Number of days to 50% flowering at M2 and M3 generations.

ND to 50% flowering reduced among the selected mutants compared to the control in the M2. Some of the mutants flowered 10-15 days earlier than the control Videza. The mutant P3N01 was first to flower at 32 DAP while the control line flowered at 47 DAP.  In the M3, further reduction in days to 50% flowering (ranged from 30 days to 36 days) was observed in M3 mutants compared to the M2. A similar result was reported by Horn (2016) in cowpea  and Shamsun et al., (2018) in chickpea. The reduction in flowering time offers the possibility for selecting early maturing mutants.
Number of days (ND) to 90% maturity at M2 and M3 generations
Fig 2 displays ND to 90% in the M2 and M3. Days to 90% maturity ranged from 50 to 66 days among the putative mutants in the M2. The earliest maturing mutants were P1N02, P3N01 and P4N03 while the latest was P10N30.  The control plant in M2 attained 90% maturity at 71 DAP. In the M3, ND to 90% maturity ranged from 48 to 60 days. The earliest to attain 90% maturity was mutant P1N02#1, while the latest were P2N09#16 and P6N10#19. The control attained 90% maturity at 70 DAP.

Fig 2: Number of days to 90% maturity at M2 and M3 generations.

It was observed that some of the mutants at the M3 matured 10-22 days earlier than the control. Reductions in days to 90% maturity due to gamma radiation treatment have been reported in many crops including cowpea (Horn, 2016) and chickpea (Shamsun et al., 2018). Early maturity is desirable for cowpea cultivation in most growing regions due to the significant loss in yield particularly when the flowering and pod filling stages coincide with long drought periods. Early maturity enables the crop to escape drought during the reproductive phase (Bowles et al., 2021). The reduction in days to 90% maturity and increased variability in yield characters proved that genetic variability induced by gamma irradiation is in the desired direction and offers an effective means for selecting early maturing and high seed yield mutants. Horn (2016) and Otusanya et al., 2022 recorded significant differences in ND to 90% maturity, 100-seed weight, seed yield per plant, number of pods per plant in M2.
Number of pods per plant (NPP) at M2 and M3 generations
NPP at the M2 and M3 are shown in Fig 3. NPP ranged from 10.00 to 45.00 among the putative mutants in M2. The highest NPP was observed in P1N05 while the lowest was recorded in P8N15. By comparison, the control lines produced a mean number of 28.90 pods per plant. In the M3, NPP ranged from 26.00 to 47.00 among selected mutants. The mutant P4N03#2 had the highest NPP (47.00) while the lowest (26.00) was observed in P1N08#13. The control line planted alongside the M3 mutants produced a mean of 30.00 pods per plant.

Fig 3: Number of pods per plant at M2 and M3 generations.

NPP increased in some of the selected mutants compared to the control in the M2. NPP increased further in the M3. Increase in NPP after mutagenic treatment has also been reported in cowpea (Horn, 2016) and chickpea (Wani, 2011).  Increase in NPP has been attributable to an increase in number of flowers in cowpea (Dingha et al., 2021). This trait plays a vital role in increased seed yield per plant in cowpea.
Number of seeds per pod (NSP) at M2 and M3 generations
NSP obtained in selected putative mutants at the M2 and M3 are presented in Fig 4. In the M2, the mean NSP varied from 10.25 to 22.50 among the putative mutants. The mutant P5N11 produced the highest NSP while P9N12 and P8N24 produced the lowest NSP. The control lines produced a mean of 12.25 of seeds per pod. NSP varied from 12.50 to 23.00 among selected mutants in M3. The highest NSP was observed in P5N07#8 while P1N06#9 had the lowest number of seeds per pod. The control lines produced a mean of 13.00 seeds per pod.

Fig 4: Number of seeds per pod at M2 and M3 generations.

NSP increased in some of the selected mutants compared to the control. Data on the mean NSP clearly showed that the mutants with the highest NSP also produced the longest pod lengths, signifying a close relationship between these two traits. Even though the mean pod length increased slightly in the M3 plants, no substantial changes in the mean NSP were observed. Similar observations were reported in soybean (Justin et al., 2012) and lentil (Laskar and Khan, 2017). Similarly, Khan and Wani (2006) did not observe significant differences in NSP in the Mand subsequent generations after gamma radiation treatments. The increase in NSP translates into high seed yielding putative mutants.
Hundred-seed weight per plant (HSWP) at M2 and M3 generations
HSWP among the selected putative mutants at the M2 and M3 are presented in Fig 5. In the M2, HSWP varied from 13.85 g to 22.80 g. The mutant P5N21 had the highest HSWP (22.80 g) while the lowest HSWP was observed in P8N24 (13.85 g).  The control line gave a mean of 16.95 g for HSWP. HSWP ranged from 12.69 g to 21.71 g in the M3.  The highest 100-seed weight was observed in P6N10#19 and the lowest was recorded in P4N14#11. The control line produced a mean of 15.28 g for HSWP.

Fig 5: 100-seed weight per plant at M2 and M3 generations.

Hundred seed weight is considered as one of the most reliable yield parameters for measuring seed yield in grain legumes. An increase in 100-seed weight was recorded in some of the selected mutants compared to the control in the M2 (from 16.95 g in the control to 22.80 g in mutant lines). A similar trend was observed in the M3 as was recorded in the M2 for HSWP. 100-seed weight increased in some of the selected mutants at M3 generation (from 15.28 g in the control to 21.71 g in mutant lines). Horn (2016) also reported significant improvement in HSWP in cowpea mutants after gamma irradiation. Similarity, Yuliasti and Reflinur (2018) reported an increase in 100-seed weight in soybean mutants. The observed increase in HSWP indicates the practicality of achieving large-seeded putative mutants via further selection.

Putative mutant lines exhibiting earliness and high seed yield at the M3 generation
Rankings of putative mutants exhibiting combined earliness and high seed yield at the M3 are presented in Table 1.  Putative mutants P1N06#9, P1N02#, P1N08#13, P1N08#17, P1N06#20, P1N08#17 P2N09#12, P4N03#2, P5N05#10 and P5N07#8 were observed to exhibit earliness as well as high seed yield. The lower the sum of ranking, the higher the position of the ranked genotypes.

Table 1: Putative mutant lines exhibiting earliness and high seed yield at M3 generation.

 Growth habit and flower colour variation
Fig 6 I and II show the growth habit and flower colour variations at the M3. In the M3, some of the induced lines changed from the semi-erect parental control growth habit to prostrate or erect growth habit in mutants (Fig 6 I). Flower colour variations at the Mare displayed in Fig 6 II. Videza produces white flowers. However, at the M3, some of the mutagenized plants produced violet and white with violet flower colours.

Fig 6: I. Growth habit variations at M3 generation.

Seed eye and coat colour variation
Seed eye and coat colour variations observed at the M3 are shown in Fig 7 I and II. Videza has black eye. Seeds with brown or brown splash eye developed at the M3 in the induced lines (Fig 7 I). Seed coat colour variations at the M3 generation are displayed in Fig 7 II. The seed coat of Videza is white. However, brown or grey speckled, brown holstein or grey Holstein coats were observed in the seeds of M3 lines.

Fig 7: I. Seed eye colour deviations at M3 generation.

Seed shape, seed texture and pod length variation
Shown in Fig 8 I and II are the seed shape and testa texture variations at the Mgeneration. Videza characteristically produces kidney shaped seeds. However, at M3, globose and Rhomboid shaped beans were obtained (Fig 8 I). Seed testa texture variations at the M3 generation are shown in Fig 8 II. Videza produced smooth to rough testa texture compared to the texture of some of the induced lines with smooth, rough to wrinkled or wrinkled seed. Pod length variations at the M3 compared to Videza are shown in Fig 9. In the M3, some of the putative mutants had pod lengths significantly longer than the control. Edematie et al., 2021 explained that it is feasible to improve seed yield in cowpea via screening for long pods.

Fig 8: I. Seed shape variations at M3 generation.


Fig 9: Pod length variations at M3 generation.

Twenty putative mutants exhibited significantly early maturity taking between 50 and 60 days to attain 90% pod maturity compared to parental control Videza lines which took 70 days. Ten mutants produced significantly higher seed yield per plant ranging 51.36 g - 97.38 g than the Videza lines with a mean of 48.65 g. Twelve putative mutant lines were ranked to outperform Videza with respect to earliness and seed yield per plant.

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