Journal of Agronomy & Agricultural Science Category: Agriculture Type: Research Article

Seed Protein Diversity Assessment and Genetic Diversity among Different Soybean [Glycine max (L.) Merrill] Accession

Thapa P1*, Dhakal K H1, Darai R2 and Shrestha A3
1 Department of genetics and plant breeding, Agriculture and Forestry University, Rampur, Chitwan, Nepal
2 National grain legumes research program, Khajura, Banke, Nepal
3 Regional agricultural research station, Parwanipur,Bara, Nepal

*Corresponding Author(s):
Thapa P
Department Of Genetics And Plant Breeding, Agriculture And Forestry University, Rampur, Chitwan, Nepal
Tel:977- 9842996262,
Email:pradip.thapa876@gmail.com

Received Date: Jun 11, 2019
Accepted Date: Jun 21, 2019
Published Date: Jun 28, 2019

Abstract

A field research was conducted at the field of National Cattle Research Program, Rampur, Chitwan from July 2015 to November 2015. The objectives of the study were to find out high protein content, high yielding and desirable traits of soybean accessions. Fifteen soybean accessions including six released varieties obtained from National Grain legume Research Program, Nepalgunj were carried out inRandomized Complete Block Design (RCBD) with three replications. No. of nodules, days to flowering, days to maturity, plant height, numbers of fruiting nodes/plant, number of branches/plant, numbers of pods/plant, numbers of seeds/plant, seed diameter, test weight and grain yield were recorded and a seed sample of 250 gm from each plot was analyzed for protein content at Department of Food Technology and Quality Control Centre, Babarmahal, Kathmandu. There was highly significant difference among the accessions in days to flowering, days to maturity, plant height, no. of fruiting nodes/plant, no. of pods/plant , no. of seeds/pod, seed diameter, test wt, grain yield and seed protein (%).Higher seed protein concentration were found in the accession PK-7394 (43.17%), LS-77-16-16 (41.33%) and PK-327 (42.11%) among fifteen soybean accessions. Accession Iang-beakong had significantly highest grain yield (3.36 ton/ha) followed by PK-7394 (3.17 ton/ha) which was at par. Accession PK-7394 was considered as the best accession in terms of both grain yield and protein content prospects.

Keywords

Correlation;Nodule;Morphology;Protein; Soybean

INTRODUCTION

Soybean [Glycine max (2n=2x=40)] is an important member of family leguminosae and sub-family papilionaceac. It is an annual herb mainly grown for seed from which oil and protein are extracted. Domestication of the soybean is believed to have originated in the northern and central regions of china as long as 5000 years ago, with the first documented use of the plant by a Chinese emperor. Soybean cultivation spread throughout Japan, Korea, and Southeast Asia, although the USA and Brazil account today for most of the soybean production of the world. Soybean primarily, an industrial crop, cultivated for oil and protein [1]. As the world population expands, there will be a greater pressure for the consumption of plant products [2]. Today soybean is considered one of the most economical and valuable agricultural commodity because of its unique chemical composition and multiple uses as food, feed and industrial materials. Soybeans have the highest protein content among cereal and other legume species, and the second highest oil content among all food legumes. Soy protein contains the essential amino acids, which closely match the requirements for humans or animals. Furthermore, soybean also contains many biological active components like is flavones, lecithin, saponins, oligosaccharides and phytosterols. Many of these components act as anti-cancer agents and antioxidants. Considering the facts there is increasing interest towards the health benefits of soy-containing foods, particularly the role of soy protein helps to lowering the incidences of certain cancers. It has been suggested that the high intakes of soy may lower incidence of certain cancers in Asian countries, where soy consumption is high, as compared to Europe or United State of America [3]. Due to its nutritional value along with its affordable low cost, soy protein is the largest commercially available vegetable protein in the world, and it is an important alternative to existing animal derived proteins. Soy proteins are also of particular interest because they impart high functionality in food systems and being used to obtain better quality products. Because of these advantages (economic, nutritive, dietetics, etc.,) it is important to develop new soy protein foods or a range of new food formulations with new textures [4].

Soybean is considered a miracle crop due to its multi-advantageous qualities i.e., food, feed, oil, fodder, soil sustainability and medicinal values. It contains about 37-42% of good quality protein, 6% ash, 29% carbohydrate and 17-24% oil comprising 85% unsaturated fatty acid with two essential fatty acids (lenoleic and linolenic acid) which are not synthesized by the human body so it is highly desirable in human diet[5].

Nitrogen is the most limiting element for crop growth and usually supplied by application of fertilizer, which brings on substantial costs to farmers and with potentially adverse effects on the environment. Being legume it also fixes atmospheric nitrogen into soil making it available to plants. As the best source of protein it truly claims the title “the meat that grows on plant”[6].Reported that soybean can capture an amount of 300 kg of nitrogen per hectare from the atmosphere. The leguminous plants establish a symbiotic relationship with rhizobia (symbiotic nitrogen fixation) to directly capture N2 to support plant growth. Nitrogen conversion takes place in a unique organ (root nodule). The development of root nodules commences with a molecular dialogue between the host plant and a compatible strain of rhizobium involving a succession of complex process that lead to profound changes in both symbiosis[7].

Latest statistics indicated that the area of soybean in Nepal was 23757 ha with an average productivity of 1.18 ton/ha (MOAD, 2015). In Nepal, soybean is being used from household consumption to commercial uses. Now soybean is considered as important crop because it is used as an important ingredient in feed industries and the soybean demands is growing rapidly. Everyday Nepal imports row or processed soybean in the estimated worth value 1-2 corer NRs from abroad (Brazil, Argentina, Ethiopia, India etc.,). With the quest of its importance, improved soybean varieties along with modern production packages are required to boost soybean production in Nepal.

The objectives of the research was
• To find out the high protein content soybean accession
• To find out the high yielding soybean accession

MATERIALS AND METHODS

The study was conducted at the research field of National Cattle Research Program, Rampur, Chitwan, from July, 2015 to November, 2015.The experiment was carried out in Randomized Completely Block Design with fifteen treatment and three replications. The plot was 8m2 (4mx2m).The experimental materials were included as six released varieties and nine pipeline accession of the soybean which were obtained from the National Grain Legume Research Program, Khajura, Nepalgunj, and BankTable 1.

S.N.

Accession name

Accession type

Origin

1

Cobb

Released variety

USA

2

Hardee

Released variety

USA

3

AGS-376

Pipeline variety

 

4

Puja

Released variety

India

5

LS-77-16-16

Pipeline variety

 

6

Seti

Release variety

Taiwan,china

7

Ransom

Released variety

USA

8

Tarkari-Bhatmas-1

Released variety

China

9

PK-7394

Pipeline variety

 

10

PK-327

Pipeline variety

 

11

PI-94159

Pipeline variety

 

12

F-778817

Pipeline variety

 

13

IARS-87-1

Pipeline variety

 

14

Iang-beakong

Pipeline variety

 

15

TGX1485-1D

Pipeline variety

 

Table 1: List of soybean accessions used for this experiment.

METHOD OF PROTEIN DETERMINATION

Sample of 250 gm from each plot was sent to the lab of Department of Food Technology and Quality Control Centre (DFTQC), Babarmahal, Kathmandu for the seed protein content calculation for which kjeldahl method is used.It is assumed, in general protein contains 16% nitrogen which means that each gram of nitrogen determined reflects a protein content of 100÷16 = 6.25g. The factor 6.25 has been worked out based on a number of studies on amino acid profile.During reporting the result, it is therefore customary to mention the factor (usually 6.25) used in the calculation. The principle for determination of nitrogen and crude protein is as follows:

A known weight of the sample was transferred to 250 ml Kjeldahl flask for determination of nitrogen by Micro-kjeldahl method. Into the flask, catalyst mixture (potassium sulphate + mercuric oxide) and concentrated H2SO4 were added. The mixture was boiled and digestion was continued until the color of the digest was colorless. The volume of the digest was made up to a known volume. Similarly a blank without the sample was run. The reduced nitrogen extracted by steam distillation from a definite volume of the digest was collected in boric acid solution. The nitrogen present in the boric acid solution was estimated by titrating with 0.02 N HCl using mixed indicator (methyl red and methylene blue). The blank distillation and titration were carried out and calculation was done.In this way nitrogen (% dry basis) is determined and finally protein content on seed is calculated by the formula
Protein(% dry basis): [Nitrogen (% dry basis) x 6.25)]

Statistical analysis

Data entry, processing and computation of mean standard deviation for all the quantitative traits were performed using Microsoft Office Excel. Data was analyzed by using R program 3.3 versions along with MS-excel 2010. Analysis of variance was performed for all traits in order to test the significance of variation among genotypes. The data was analyzed for mean, coefficient of variation (CV %), LSD value and correlation coefficient. UPGMA clustering was done using Minitab 14.

Cluster analysis

Cluster analysis is a type of multivariate technique whose primary purpose is to group individuals or objects based on characteristics they possess, so that individuals with similar description are mathematically gathered into the same cluster. The resulting cluster of individuals should exhibit higher within clusters homogeneity and between clusters heterogeneity. Thus, if the classification is successful, individuals within cluster should be closer when plotted geometrically and different clusters shall be apart. Among various agglomerative hierarchical methods, Unweighted Paired Group Method using Arithmetic Averages (UPGMA) is most commonly adopted clustering algorithm, followed by the Ward’s minimum variance method (Ward, 1963).The accessions were clustered using days to flowering, days to maturity, plant height,no of fruiting nodes/plant, pods/plant, seeds/pod, seed diameter, grain yield and protein concentration as variables.

RESULTS AND DISCUSSIONS

There was variation found in agro-morphological characters among the fifteen soybean accession shown in Table 2. Variation was found mainly in hypocotyl colors, leaflet shape, pubescence density, pubescence color, flower color, seed coat color, hilum color, surface luster and plant height. Two types of hypocotyls color (green and purple) were recorded. Among the fifteen soybean accessions, eight soybeans had green hypocotyl color and remaining seven had purple hypocotyl .Three different types of leaflets shape were observed. Among them; eleven soybean accessions had intermediate, two accessions had broad and remaining two accessions had narrow type of leaflets shape. Variation wasalso observed in pubescence color and density too. Among the tested accessions, eight accessions had normal, three had semi-sparse, three had dense and one accession had sparse type of pubescence density. Tawny and grey color of pubescence was observed in eleven and four accessions respectively. Among the fifteen accessions; nine and six accessions had white and purple flower respectively. Three types of seed coat color were observed among the tested accession. Ten accessions had yellow, three accessions had yellowish white and remaining one accession i.e. Tarkari-Bhatmas-1 had green type of seed coat color. Brown, black and grey hilum color was found in six, five and four accessions respectively. Surface lusture was also varied, thirteen accessions had intermediate type of surface lusture and remaining (Hardee and Iang-beakong) accessions had shiny type of surface lusture. There was significant variation found.In plant height also. Significantly higher plant height (57.80 cm) was recorded in Hardee accession and lowest plant height (32.97cm) was recorded in Tarkari-bhatmas-1.
 

S.N

Genotype Name

Hypocotyl color

Leaflets Shape

PubescenceDensity

PubescenceColor

Flower Color

Seed coat color

Hilum color

Surface lusture

Pht(cm)

 
 

1

Cobb

Green

Intermediate

Dense

Brown

White

Yellow

Brown

Intermediate

45.37

 

2

Hardee

Green

Intermediate

Dense

Brown

White

Yellow

Brown

Shiny

57.8

 

3

AGS-376

Green

Broad

Dense

Brown

White

Yellow

Black

Intermediate

52.56

 

4

Puja

Green

Intermediate

Normal

Brown

White

Yellow

Black

Intermediate

43.12

 

5

LS-77-16-16

Purple

Narrow

Normal

Grey

Purple

Yellowish white

Grey

Intermediate

26.79

 

6

Seti

Green

Intermediate

Normal

Brown

White

White

Brown

Intermediate

41.85

 

7

Ransom

Purple

Intermediate

Normal

Brown

Purple

Yellow

Black

Intermediate

40.54

 

8

Tarkari-Bhatmas-1

Purple

Intermediate

Normal

Grey

 Purple

Green

Black

Intermediate

32.97

 

9

PK-7394

Purple

Narrrow

Sparse

Brown

Purple

Yellowish white

Brown

Intermediate

54.84

 

10

PK-327

Purple

Intermediate

Semi Sparse

Grey

Purple

Yellowish white

Grey

Intermediate

27.89

 

11

PI-94159

Green

Intermediate

Normal

Brown

White

Yellow

Brown

Intermediate

37.13

 

12

F-778817

Green

Intermediate

Semi Sparse

Brown

White

Yellow

Grey

Intermediate

35.25

 

13

IARS-87-1

Green

Broad

Semi Sparse

Brown

White

Yellow

Black

Intermediate

39.99

 

14

Iang-beakong

Purple

Intermediate

Normal

Brown

Purple

Yellow

Brown

Shiny

57.05

 

15

TGX1485-1D

Purple

Intermediate

Normal

Grey

White

Yellow

Grey

Intermediate

47.1

 
 
 

Table 2:Agro-morphological characteristics of 15 soybean accessions.

Agronomic Performances of Soybean Accession

Analysis of Variance (ANOVA) revealed that there was significant difference (P ≤ 0.01) in days to flowering, Days to maturity, Plant height, nodes/plant, pods/plant, seeds/pod, seed diameter, grain yield and seedprotein content [8]. also reported range of variation for days to flowering, days to maturity, plant height, no of pods/plant, 100 seed weight and grain yield.There was no significant difference in nodule number among the accession. Significantly longer days to flowering are observed in Iang-beakong accession (50 days) and significantly shorter days to flowering are observed in PI-94159, PK-327, LS771616, Ransom and Tarkari-bhatmas-1. Similarly AGS-376 accession is recorded as a late mature (127 days) and Tarkari-Bhatmas accession is earlier maturity (94 days). Result showed that Hardee and IangBeakong accession had significantly higher plant height and LS-771616 had lowest plant height Significantly higher number of pods/plant was observed in IangBeakong accession (72 pods/plant) and significantly lower number of pods/plant was observed in cobb accession (30 pods/plant). Soybean plant with higher plant height and higher no of nodes might bear higher no of pods/plant. Therefore genotypes with remarkably higher number of pods/plant can be utilized in hybridization of soybean with early flowering and maturing traits for better yield [9,10].Significantly greater seed diameter wasrecorded in IARS-87-1 accession (5.38mm) and significantly lower seed diameter wasobserved in LS-771616 accession (4.39mm). IangBeakong accession showed highest yield i.e. 3.38 ton/ha among the accession and Cobb showed significantly lower yield i.e., 1.72 ton/ha. Result showed that PK-7394 accession had significantly higher protein content (43.17%) among the fifteen accession which was statistically at par with protein content of PK-327 (42.11%) and LS-771616 (41.93%). Substantial genotypic variation for seed protein concentration has been documented by [11-16]. But high seed protein concentration is frequently associated with less yield [17-19] Table 3.

Genotype

DF

DM

Nodnm

Pht(cm)

Nodes

Branch

Pods

Seeds

TW

SD(mm)

GY

Protein (%)

Cobb

42

125

48

45.37

11

4

30

1.93

143

5.12

1.72

39.73

Hardee

45

126

53

57.8

13

5

59

1.9

123

4.87

2.9

41.13

AGS-376

43

127

55

52.56

10

5

54

2.03

133

4.86

2.8

40

Puja

42

117

44

43.12

9

4

40

2

140

4.99

2.2

36.66

LS-77-16-16

40

100

45

26.79

9

4

44

2.26

100

4.39

1.76

41.93

Seti

40

101

38

41.85

11

3

39

1.8

130

5.05

2.03

41.65

Ransom

40

103

55

40.54

10

3

37

2.03

140

4.98

2.07

37.94

Tarkari-Bhatmas-1

40

94

42

32.97

9

2

35

1.96

153

5.17

2.15

35

PK-7394

50

110

50

54.84

12

3

70

1.96

113

4.76

3.17

43.17

PK-327

40

101

47

27.89

9

4

48

2.3

105

4.39

2.02

42.11

PI-94159

40

116

38

37.13

9

4

44

2.03

130

5.13

2.29

38.11

F-778817

43

117

47

35.25

9

4

42

2.16

153

5.1

2.58

35.93

IARS-87-1

43

117

40

39.99

11

4

49

2.23

157

5.38

3.07

39.65

Iang-beakong

50

112

42

57.05

13

4

72

1.96

117

4.89

3.36

34.85

TGX1485-1D

43

117

52

47.1

11

3

42

2.03

127

5.08

2.13

40.07

Mean

42.73

112.24

46.37

42.68

10.46

3.69

46.95

2.04

130.89

4.94

2.42

39.19

P value

0

0

0.89

0

0.0063

0.0995

0.01

0

0

0.001

0

0

F value

7.13*1029

896.71

0.52

8.88

3.006

1.7563

2.74

4.1317

9.93

3.92

7.55

31.35

CV%

1.60E-14

0.53

26.12

13.48

17.96

29.65

33.98

8.13

7.35

4.78

1.53E-14

2.1

LSD0.05

1.14E-14

0.99

20.26

9.62

3.14

1.83

26.69

0.27

16.1

0.39

6.17E-16

1.38

Table 3: Agronomical performance of fifteen soybean accessions.
Notes: DF=Days to flowering,DM=Days to maturity,Nodnum=Nodule Number,Pht=Plant height,TW=Test Weight,SD=Seed Diameter,GY=Grain Yield

Correlation of different traits of soybean with protein content

Protein content was significantly and negatively associated with seed diameter and test weight(Table 4). Similar result was showed by Mario et al., (2000). According to him, the negative correlation between seed diameter and protein content % was relatively low, although significant (-0.42) and Burton(1991) reported an estimate of correlation coefficient as high as -0.47 and Johansonet al., (1955) reported an estimate of -0.64 correlation coefficient between protein content and grain yield.Tiniuset al.,(1993) reportedcorrelationvalues for seed size and protein content in two soybean sub-populations as -0.97 and -0.86.There was non-significant negative correlation between protein content and grain yield found in our studywhich might be due to influence of environmental factors during the research period. A decrease in seed protein content associated with deficit stress late in the season may have been caused by a concurrent increase in temperature or decrease in nitrogen supply.

Characters

Nodnum

GY(ton/ha)

SD(cm)

Test wt(gm)

Protein(%)

Nodnum

1

0.030ns

-0.155ns

-0.023ns

0.067ns

GY(ton/ha)

 

1

0.129ns

0.023ns

-0.117ns

SD(cm)

 

 

1

0.740**

-0.35*

Test wt (gm

 

 

 

1

-0.479**

Protein(%)

 

 

 

 

1

Table 4: Correlation of different traits with protein concentration.
Notes: *= Significant & ns= Non-significant

According to previous finding correlation between seed protein content and grain yieldwas negative[13,18,20]. The negative correlation of protein content of seed with grain yield, seed diameter and test weight indicated that it would be very difficult to identify a soybean accession having higher grain yield simultaneously with higher protein content. Increase in one trait would result in reduction of the other, that is simultaneously increase or decrease of both traits would be difficult. Breeding for high seed protein concentration was based on increase in assimilates supply per seed. This increase however was more related with less seed set than more leaf area. This might be the link between high protein concentration and lower yield.

Nodule number was positively correlated with grain yield and protein content (Table 3).It might be due to nitrogen fixation process by the help of nodule present in roots of soybean plant. Due to the nitrogen fixation process, nitrogen content in the soil may increase and ultimately increase in yield also.

UPGMA clustering

The accessions were clustered using days to flowering, days to maturity, plant height,no of fruiting nodes/plant, pods/plant, seeds/pod, seed diameter, grain yield and protein concentration as variables. The dendrogram is presented in Figure 1.The critical examination of the dendrogram revealed five clusters with minimum of 29.35% similarity level in UPGMA clustering. Clusters were obtained on the basis of similarity percentage and related characters.

Figure 1: UPGMA clustering of 15 soybean accessions.

Cluster 1 consisted of one soybean accession i.e. Cobb. This cluster had lowest no of pods/plant and grain yield.Cluster 2 consisted of two soybean accession. It includes Hardee and AGS-376.This cluster contained late matured accession which had higher plant height and higher concentration of protein and relatively higher grain yield.Cluster 3 consisted of five soybean accessions. It include Puja, TGX1485-1D, PI-14159,F-778817 andIARS-87-1.These soybean accessions had intermediate value for days to flowering, days to maturity, plant height, no of pods/plant, grain yield and protein concentration.

Cluster 4 consisted of five soybean accessions. It include LS-77-16-16, PK-327, Seti, Ransom and Tarkari-Bhatmas-1.These were early matured accessions which had lowest plant height, no of fruiting nodes/plant and seed diameter. These accessions had intermediate value for protein concentration and grain yield. Cluster 5 consisted of two soybean accessions including PK-7394 and Iang beakong. These accessions had longer days to flowering, highest plant height, pods/plant, nodes/plant as well as grain yield among the accessions and intermediate value for the protein concentration.

CONCLUSION

High protein content is an important specialty trait for soybean breeding programs. Soybean is an important ingredient for livestock, poultry and other industry and contributes more than 70% of the protein consumed by humans. In this studyPK-7394 (43.17%), LS-77-16-16 (41.33%) and PK-327 (42.11%) were identified as high protein content accessions. Among the studied accessions, Iang-beakong had significantly higher grain yield(3.36 ton/ha) followed by PK-7394 (3.17 ton/ha).In the correlation analysis, grain yield showed positive and highly significant association with days to 50% flowering, days to maturity, plant height, no of nodes/plant. These traits can be considered for selection programs aimed for yield improvement. In terms of both protein content and grain yield,PK-7394 accession was found superior at Rampur locality.

ACKNOWLEDGEMENT

I am grateful to acknowledge National Agricultural Research and Development Fund (NARDF), Singh Durbar Plaza, Kathmandu for providing the required fund during my research works. I would also like to thank Department of Food Technology and Quality Control, Kathmandu for timely help in the protein content analysis.

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Citation: Thapa P, Dhakal K H, Darai R, Shrestha A (2019) Seed Protein Diversity Assessment and Genetic Diversity among Different Soybean [Glycine max (L.) Merrill] Accession. J Agron Agri Sci 2: 009.

Copyright: © 2019  Thapa P, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


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