Nghiên cứu cải tiến các giống đậu tương DT2008, DT96 và đt26 bằng phương pháp xử lý đột biến tia gamma nguồn co60 tt tiếng anh

MINISTRY OF EDUCATION AND TRAINING MINISTRY OF AGRICULTURE AND RURAL DEVELOPMENT VIETNAM ACADEMY OF ARICULTURAL SCIENCES ---*--- NGUYEN VAN MANH RESEARCH ON IMPROVING SOYBEAN VARIETIE

MINISTRY OF EDUCATION

AND TRAINING

MINISTRY OF AGRICULTURE AND RURAL DEVELOPMENT VIETNAM ACADEMY OF ARICULTURAL SCIENCES

-* -

NGUYEN VAN MANH

RESEARCH ON IMPROVING SOYBEAN VARIETIES DT2008, DT96 AND ĐT26 THROUGH 60 Co GAMMA IRRIGATION

Research: Genetics and Crop breeding

Code: 9620111

DOCTORAL THESIS SUMMARY OF AGRICULTURE

Hanoi - 2020

The Doctoral thesis was completed in:

VIETNAM ACADEMY OF AGRICULTURAL SCIENCES

PhD thesi can be found at:

1 National Library of Vietnam

2 Library of Vietnam Academy of Agricultural Sciences

INTRODUCTION

1 Imperativeness of the thesis

Soybean (Glycine max (L.) Merrill), a short-day crop, has high nutrient

content (Protein 30 - 45%, lipit 18 – 22% ), easy cultivation, wide adaptation, effect on land reclamation and insect limitation in the rotation of other crops

In Vietnam, soybean is a traditional crop and grown in 28 of 63 provinces/cities It plays an important role in agricultural production and has high economic value and efficiency in crop restructing but its area is decreasing In 2018, soybean production reached at only 53.1 thounsand hecta with the productivity of 80.8 thounsand tons (https://www.gso.gov.vn), only met about 8% of domestic demand, the rest had to imported The causes

of decrease in soybean production are the lack of soybean varieties with high yield, good quality and good resistance; difficult condition of cultivation; backward cultivation techniques and low intensive investment Therefore, it

is urgent and necessary to create new soybean varieties with high yield, good quality and suitable for many different crops In soybean, it is hard to eliminate anthers because its flowers are small, perishable and vulnerable In addition, its completely self-fertilization results in being difficult to create heritable variations by hybridization

Therefore, mutation method through 60Co gamma irradiation is so efficient to generate new heritable variations in soybean, contributing to diversifying genetic variations, shortening the time to select a new variety, and especially improving existing soybean varieties In order to diversity soybean varieties in production, contributing to expanding the area and developing soybean production in Vietnam, the thesis “Research on improving soybean varieties DT2008, DT96 and ĐT26 through 60Co gamma irradiation” has been carried out

2 Objectives

Improving soybean varieties in the goals of shortening the growth duration and stem height, increasing the resistance to lodging and seed quality…through 60Co gamma irradiation to create and select new soybean varieties, meeting the demand of current production

3 Scientific and Practical Significances

3.1 Scientific Significances

Results of the thesis would be the basic for creating new soybean varieties by mutation method through 60Co gamma irradiation in the goals of shortening the growth duration and stem height, increasing the resistance to lodging and seed quality…

Through conducting the thesis, the highly effective irradiation doses

of gamma ray (60Co) have been identified for creating new mutant soybean varieties

The data published in this work can be used as a guide material for teaching and research on soybean breeding in agricultural universities and research institutes

3.2 Practical Significances

New soybean mutants including 01 self- declared circulation variety DT215 and 02 promising lines DT96ĐB and ĐT26ĐB have been created They have high yield (>2.5 tons/ha), good quality (protein > 40%, lipid

>18%), good resistance to rust and powder mildew, meeting the demand of current production

The group of 234 mutant soybean lines in the M7 generation divided from soybean virieties DT2008, DT96 and ĐT26 with improved traits are signigicant materials for creating and selectiong new soybean varieties

4 Research Materials and Scope

4.1 Research Materials

Soybean varieties DT2008, DT96 and ĐT26

Mutagen of gamma ray from the source of Cobalt 60 (60Co)

A collection of 234 mutant soybean lines with one or some improved traits compared to the original varieties such as growth duration, stem height, seed cover colour… have been created and used as significant materials for new soybean breeding

A new mutant soybean variety DT215 (the yield of 2.71 – 3.32 tons/ha but black seed coat different from DT2008’s yellow seed coat, 5 – 10 days shorter growth duration in the range of 90 – 95 days) which was self-recognized for introducing into the production and 2 promising mutant

soybean varieties DT96ĐB (the growth duration of 90 – 94 days, the height

of 52.1 – 59.9 cm and the yield of 2.22 – 3.06 tons/ha but better lodging resistance compared to the origin DT96) and ĐT26ĐB (the yield of 2.23 – 2.55 tons/ha but black seed coat different from ĐT26’s yellow seed coat, the growth duration of 86 – 91 days) have been produced They have good growth and development, good resistance to deseases, high yield… and satisfy the demand of current produciton

6 Thesis structure

The thesis consists of 140 pages (excluding illustrations and Appendices) including Introduction (4 pages); Chapter 1: Literature Review (35 pages); Chapter 2: Materials, Contents and Research Methods (12 pages); Chapter 3: Research Results and Discussion (68 pages); Conclusions and Recommendations (2 pages); List of published works related to the thesis (1 page); References (18 pages) with 85 Vietnamese documents, 49 English documents and 3 websites The thesis has 53 data tables, 12 pictures, 35 appendices and 6 published works

CHAPTER I LITERATURE REVIEW

The literature review presents the following contents of (1) Introduction

of soybean, (2) Research situation of soybean breeding in the world and in Vietnam, (3) Mutaiton and applications in plant breeding, (4) Research on mutant soybean breeding Thereby, some following comments have been drawn: Soybean is the main source of protein and vegetable oil for the whole world It is used as food for human and livestocks, for soil conditioning… In Vietnam, soybean is a traditional crop but its production has met only about 8%

of domestic demand with the decrease in area because of low yield, backward cultivation techniques, low investment, especially the lack of new soybean varieties…

Mutation method is so effective to improve plant varieties and has created over 3,200 varieties of 220 plant species It has produced a rich source of genetic variation, especially unavailabe traits in nature In Vietnam, there has been 61 mutant crop varieties of rice, flower, soybean…

Currently, there has been 171 mutant soybean varieties in the world, mainly created through gamma irradiation (66 varieties) with the improved traits

of yield, growth duration, stem height, seed coat colour and resistance In Vietnam, there has been 11 mutant soybean varieties, of which 8 varieties created through gamma irradiation such as DT84, DT90, DT99, DT2008, AK06… However, these varieties has been mainly created through gamma irradiation on dry seeds There has been still the limitation on studies of gamma irradiation on germination seeds and flowering plants have been still limited

CHAPTER II

MATERIALS, CONTENTS AND RESEARCH METHODS 2.1 Materials

2.2.1 Soybean varieties: DT2008, DT96, ĐT26 and DT84 (control)

- Irradiation doses on dry seeds: 0 (control), 100, 150, 200, 250, 300 and

growth and development of soybean varieties

Effect of gamma irradiation (60Co) on the growth and development of soybean varieties

Effect of gamma irradiation (60Co) on variation frequency and spectrum

Evaluating and comparing promising mutant soybean lines

Evaluating the genetic diversity of some mutant soybean lines by SSR markers

Content 3: Testing the production trials of promising mutant soybean

varieties

2.3 Research Methods

2.3.1 Gamma irradiation method soybean varieties DT2008, DT96 and ĐT26

a) Gamma irradiation on dry seeds

Dry seeds of soybean varieties were irradiated at the doses of 0 (control),

100, 150, 200, 250, 300 and 350 Gy with the source power of 64.8 kCi during

30 minutes Each treatment used 600 seeds

b) Gamma irradiation on germination seeds

Dry seeds of soybean varieties were soaked in fresh water during 2 hours, and then put into the incubator at 260C during 2, 4, 6, 8 and 10 hours The seeds after incubating were irradiated at t0he doses of 0 (control), 25, 50, 75, 100,

125 and 150 Gy during 30 minutes with the source power of 62.3 kCi Each

treatment used 500 seeds

c) Gamma irradiation on flowering plants

Dry seeds of soybean varieties were sown in pots At the blooming stage (50% of plants in bloom), flowers, pods and buds on each plant of pots were marked with thread separately and irradiated at the doses of 0 (control), 10, 20,

30, 40, 50 and 60 Gy during 30 minutes, the source power of 61.1 kCi Each treatment used 3 pots

development of soybean varieties

a) Experimental design

Treatments were designed in sequence without replication and with the alternation of the controls of original varieties The plot area was 8.5 m2 (5m x 1.7m) Seeds were sown in the density of 35-40cm in row distance and 10cm in hole distance, one seed per hole

* M1 generation:

Gamma irradiation on dry seeds: After irradiaton, 100 seeds of each treatment were taken and sown on the tray with sterilized sand at 26oC during 7 days to evaluate the germination rate The remaining seeds of each treatment were sown in the experimental field to evaluate the growth and development At the harvesting stage, variant individuals were collected separately The mixture

of remaning plants were collected in each treatment

Gamma irradiation on germination seeds: Seeds after irradiation were sown

in the experimental field to evaluate the growth and development At the harvesting stage, variant individuals were collected separately The mixture of remaning plants were collected in each treatment

Gamma irradiation on flowering plants: After irradiaton, soybean varieties were observated and evaluated on the growth and development in green house At harvesting stage, seeds from marked flowers, pods and buds were collected separately in each treatment

d) Effect of gamma irradiation ( 60 Co) on variation frequency and spectrum of soybean varieties

Observating and detecting morphological variations and agronomic traits such as leaflet shape, stem colour, seed coat colour, growth duration, stem height…

e) Evaluating the inheritance of morphological variations derived from soybean varieties irradiated with gamma ray ( 60 Co)

The inheritance of morphological variations was evaluated and monitored over generations M1 to M2, M2 to M3 and M3 to M4

2.3.3 Research on selecting significant mutant soybean lines for soybean breeding

a) Research on selecting significant mutant soybean lines for soybean breeding

Pedigree method was used to select mutant soybean lines based on agrobiological traits such as stem shape, leaflet shap, pubescence colour, growth duration, lodging resistance, 1000-seed weight, seed coat colour…

bio-b) Evaluating and comparing promising mutant soybean lines

Promising mutant soybean lines were evaluated and compared under the national standard QCVN 01-58:2011/BNNPTNT by Ministry of Agriculture and Rural Development on the value of cultivation and use of soybean varieties Experiments of comparing mutant soybean lines were designed in RCB with three replications on the plots with the area of 8.5 m2

c) Evaluating the genetic diversity of some mutant soybean lines by SSR markers

2.3.4 Testing the production trials of promising mutant soybean varieties

Promising mutant soybean varieties were tested under the national standard QCVN 01-58:2011/BNNPTNT by Ministry of Agriculture and Rural Development on the value of cultivation and use of soybean varieties

2.3.5 Analysis of grain nutrition content

2.3.6 Observation traits

Observation traits and evaluation methods under the national standard QCVN 01-58:2011/BNNPTNT by Ministry of Agriculture and Rural Development on the value of cultivation and use of soybean varieties

2.3.7 Data Analysis

Data analysis was processed by Excel 2016 and IRRISTAT 5.0

2.4 Research time and place

CHAPTER III RESEARCH RESULTS AND DICUSSIONS 3.1 Research on the effect of gamma irradiation ( 60 Co) on the growth and development of soybean varieties

development of soybean varieties irradiated on dry seeds

a) Effect of gamma irradiation ( 60 Co) on germination and survival rate of soybean varieties irradiated on dry seeds

There was a little effect of gamma irradiation on germination rate in

M1 and M2 generations in soybean varieties irradiated on dry seeds The germination rate ranged from 98 to 100% (the rate of 100% at the control) in

M1 generation and from 98.3 to 100% (the rate of 100% at the control) in M2generation

Table 3.1 Effect of gamma irradiation (60Co) on germination rate of soybean

varieties in M1 and M2 generations

Unit: %

Radiation

doses

M1 generation M2 generation DT2008 DT96 ĐT26 DT2008 DT96 ĐT26

Table 3.2 Effect of gamma irradiation (60Co) on survival rate of soybean varieties irradiated on dry seeds in M1 and M2 generations

Unit: %

Radiation

doses

M1 generation M2 generation DT2008 DT96 ĐT26 DT2008 DT96 ĐT26 0Gy (control) 97.6 94.6 96.4 85.2 80.6 83.1

c) ) Effect of gamma irradiation ( 60 Co) on variation frequency and spectrum of soybean varieties irradiated on dry seeds

Figure 3.1 Effect of gamma irradiation (60Co) on variation frequency of soybean varieties irradiated on dry seeds in M1 and M2 generations Variation frequency was on the increase of radiation doses from 100

Gy to 350 Gy In M1 generation, variation frequency of soybean varieties DT2008, DT96 and ĐT26 ranged from 0.6 – 58.8% (the rate of 0.4% at the control), 0.2 – 54.6% (the rate of 0.2% at the control) and 0.4 – 52.6% (the rate of 0.2% at the control) respectively In M2 generation, variation frequency of soybean varieties DT2008, DT96 and ĐT26 ranged from 6.1 – 36.1% (the rate of 0.8% at the control), 6.4 – 39.8% (the rate of 0.6% at the control) and 6.4 – 37.7% (the rate of 0.5% at the control) respectively

0 (Đ/c) 100 150 200 250 300 350

Table 3.10 Effect of gamma irradiation (60Co) on variation spectrum of soybean varieties irradiated on dry seeds in M1 and M2 generations

Unit: number of variations

Radiation

doses

M1 generation M2 generation DT2008 DT96 ĐT26 DT2008 DT96 ĐT26

Gamma irradiation on dry seeds produced many variations in M1 and

M2 generations such as stem shape (curved or flat stem…), branch shape (early branching, symmetrical branching…), leaflet shape, seed coat colour, pubescence colour… Variation spectrum was on the increase with increasing doses from 100 Gy to 200 Gy and on the decrease at the dose of 300 Gy In

M1 generation, the number of variations of soybean varieties DT2008, DT96 and ĐT26 ranged from 2-9, 1-9 and 2-9 respectively In M2 generation, the number of variations of soybean varieties DT2008, DT96 and ĐT26 was more than in M1 generation and ranged from 2-14, 3-14 and 2-14 respectively The radiation dose with the most number of variations was 200

Gy (9 variations in M1 generation and 14 variations in M2 generation), followed by the doses 250 Gy (7-8 variations in M1 generation and 10-14 variations in M2 generation) and 150 Gy (6-7 variations in M1 generation and 7-14 variations in M2 generation) There were a little number of variations at the doses 100, 300 and 350 Gy In particular, there were some significant variations for soybean breeding in M2 generation such as straight stem (acute-angled branching angle), branchiness, black seed cover, fruitfulness, early maturation…

development of soybean varieties irradiated on germination seeds

a) Effect of gamma irradiation ( 60 Co) on germination and survival rate of

soybean varieties irradiated on germination seeds

In M2 generation, the germination rate of soybean varieties DT2008, DT96 and ĐT26 at treatments was equal to the control, ranged from 98.7-

99.7% (the rate of 99.5% at the control), 98.7-99.5% (the rate of 99.8% at the control) and 98.7-99.7% (the rate of 99.9% at the control)

Table 3.12 Effect of gamma irradiation (60Co) on germination rate of soybean

varieties in M1 and M2 generations

Unit: %

Radiation

doses

M1 generation M2 generation DT2008 DT96 ĐT26 DT2008 DT96 ĐT26 0Gy (control) 96.2 92.4 95.1 84.8 82.1 83.2

The survival rate of soybean varieties at treatments was lower than that

at the control and on the decrease with the increase of radiation doses from

25 Gy to 150 Gy, and was the lowest at 150 Gy The effect of gamma irradiation on the survial rate in M1 generation was higher than in M2generation In M1 generation, the survival rate of soybean varieties DT2008, DT96 and ĐT26 ranged from 12.3 – 91.4% (the rate of 96.2% at the control), 8.1 – 87.4% (the rate of 92.4% at the control) and 10.8 – 90.0% (the rate of 5.1% at the control) respectively Lethal dose 50% (LD50) was indentified at

100 Gy In M2 generation, the survival rate of soybean varieties DT2008, DT96 and ĐT26 ranged from 78.0 – 83.7% (the rate of 84.8% at the control), 74.9 – 81.7% (the rate of 82.1% at the control) and 74.8 – 81.0% (the rate of 83.2% at the control) There was a little effect of different incubation times

on the survival rate

b) Effect of gamma irradiation ( 60 Co) on growth and development indicators of soybean varieties irradiated on germination seeds

Gamma irradiation on germination seeds prolonged the growth duration

and reduced quantity traits such as stem height, a number of fertile pods, individual yield in soybean varieties The higher the dose of gamma irradiation was, the greater the effect of gamma irradiation on growth and development was in M1 generation There was no difference of growth and development among treatments and the control in M2 generation

c) Effect of gamma irradiation ( 60 Co) on variation frequency and spectrum of soybean varieties irradiated on germination seeds

Variation frequency of soybean varieties was on the increase with the rise in gamma irradiation dose In M1 generation, variation frequency of soybean varieties DT2008, DT96 and ĐT26 ranged from 24.1 – 100, 25.3 – 100.0 and 20.4 – 100% (the rate of 0% at the control) respectively From 100Gy and over doses, variation frequency was 100% (mainly sterile variations and late maturation) In M2 generation, variation frequency of soybean varieties DT2008, DT96 and ĐT26 ranged from 10.3 – 58.1, 23.9 – 88.0 and 16.9 – 68.7% respectively There was a little effect of different incubation times on the variation frequency in soybean varieties

Figure 3.2 Effect of gamma irradiation (60Co) on variation frequency of soybean varieties irradiated on germination seeds in M1 and M2 generations Table 3.18 Effect of gamma irradiation (60Co) on variation spectrum of soybean varieties irradiated on dry seeds in M1 and M2 generations

Unit: number of variations

Radiation

doses

M1 generation M2 generation DT2008 DT96 ĐT26 DT2008 DT96 ĐT26