Evaluation of salt tolerance of some local rice varieties in the Mekong Delta.. Evaluation of salt tolerance of 12 local rice varieties in Tra Vinh province by DNA markers and K+/Na+...
Trang 1MINISTRY OF EDUCATION AND TRAINING
CANTHO UNIVERSITY
SUMMARY OF DISERTATION
SPECIALIZATION: CROP SCIENCES
Code: 62 62 01 10
TRAN HUU PHUC
COLLECTION, SELECTION AND PURE LINES SELECTION OF SALT-TOLERANT RICE VARIETIES FOR RICE-SHRIMP
SYSTEMS
Cantho, 2021
Trang 2THE THESIS HAS BEEN CONDUCTED IN
1streviewer: Dr TRAN NGOC THACH
2ndreviewer: Assoc Dr TAT ANH THU
Dissertation can be further referred at:
1 Vietnam National Library
2 Learning Resource Center- Cantho University
Trang 3LIST OF PUBLISHED JOURNAL
International Journal
1) Tran Huu Phuc, Van Quoc Giang, Nguyen Van Manh,
Huynh Ky, 2021 Genetic diversity of local rice varieties (Oryza sativa L.) in Vietnam’s Mekong Delta based on SSR markers and morphological characteristics Indonesian Journal of Biotechnology, Volume 26(2), 2021, 76‐81
Domestic Journal
1) Tran Huu Phuc, Vu Anh Phap, Huynh Ky and Van
Quoc Giang, 2019 Collection, selection and pure lines selection of
Ba bong man and Bo liep 2 Vietnam Agricultural Science Journal, Volume 2(99)/2019
2) Tran Huu Phuc, Vu Anh Phap, Nguyen Lam Minh, Tran
Thi Xuan Mai and Pham Van Mich, 2018 Identification and assessment of salt tolerance in rice varieties based on Simple Sequence Repeats (SSR) markers Can Tho University Journal of Science, Volume 6B (2018): 82-89
3) Huynh Ky, Tran Huu Phuc, Van Quoc Giang, Nguyen
Van Manh, Nguyen Loc Hien and Nguyen Chau Thanh Tung, 2019 Selection of high quality rice varieties using functional Wx gene Vietnam Agricultural Science Journal, Volume 8(105)/2019
4) Huynh Ky, Tran Huu Phuc, Van Quoc Giang, Tran Thi
Yen Nhi, Nguyen Loc Hien, Nguyen Chau Thanh Tung, 2019 Evaluation of salt tolerance of some local rice varieties in the Mekong Delta Vietnam Agricultural Science Journal, Volume 7(104)/2019
5) Huynh Ky, Van Quoc Giang, Nguyen Chau Thanh Tung,
Nguyen Loc Hien and Tran Huu Phuc, 2018 Evaluation of salt
tolerance of 12 local rice varieties in Tra Vinh province by DNA markers and K+/Na+ Can Tho University Journal of Science, Volume 9B (2018): 41-46
Trang 4CHAPTER 1: INTRODUCTION 1.1 Problem statement
Local rice plants have grown for a long time in acid and saline soils Local rice plants have good tolerance to harsh farming conditions as evidenced by salt tolerance, drought tolerance, and waterlogging tolerance Before the 1960s, the whole Mekong Delta was cultivated with local rice varieties that were distributed according to different ecological conditions, such as floating rice in flooded areas >1 m, deep-water rice for 0.5–1.0 m inundation, and salt-tolerant local rice in inundated coastal areas <50 cm This adaptation led to thousands of rice varieties being cultivated in the Mekong Delta Today, the local rice area is greatly reduced, so the diversity
in the number of varieties has decreased significantly and faces the risk of extinction The source of seeds grown in the field is mixed with different heights and morphological characteristics, and the quality of commercial rice
is decreasing because local rice varieties are rarely purified Some areas in the Mekong Delta also grow local rice where it is not possible to meet the requirements of intensive farming or where the farming conditions are harsh
In areas that primarily use rain water, the land has not been properly improved and the investment level of farmers is still very limited Local rice plants are the main crop for the coastal acid sulfate soils with rice-shrimp farming and rainwater-dependent land
In recent years, local rice plants have been grown in the rice-shrimp system and rice-fish system because the local rice has good tolerance to the saline soil conditions of the Mekong Delta Focusing on finding and developing local rice varieties suitable for today's harsh weather conditions
is very important The goal is to provide seed resources for areas with difficult farming conditions This contributes to preserving the precious genetic resources of traditional local rice and contributes to the recovery of local "specialty" plant varieties Finding and recovering local rice varieties to put into production will contribute to improving the yield and quality of local rice varieties, thus increasing people's incomes and ensuring food security This study is thus proposed for implementation
Trang 51.2 Objectives
The objective is the collection and evaluation of some local rice varieties in some coastal provinces in the Mekong Delta The aim is to identify two purified local rice varieties that exhibit salt tolerance, suitability for rice-shrimp systems, high yield (average ≥3.5 tons/ha), chalkiness of endosperm ≤15%, amylose ≤24%, and genes for salt tolerance and BPH resistance
1.3 Scientific significance
Some local rice varieties have also been collected in saline areas
of the Mekong Delta, contributing to the conservation of local rice varieties This has contributed to research and teaching
The application of molecular markers in the study of quality, salt tolerance, and brown planthopper resistance has contributed to increasing the accuracy of the research The data in this study can contribute to important teaching and reference work
1.4 Practical significance and applicability of the thesis
The study successfully selected and purified two local rice varieties, contributing to providing a source of salt-tolerant local rice varieties It has provided a source of high-quality seed for rice production
in the rice-shrimp model
1.5 New contributions of the thesis
The data obtained from the thesis provides a part of the data set,
to build a database of 41 local rice genetic resources as well as a database
of rice genetic resources; The research results of the thesis have provided more information for breeders about the morphological, agronomic, and salt tolerance characteristics of 41 local rice varieties; The research results
of the thesis also contribute to the practice of two local rice varieties that are pure-filtered: Ba bong man and Bo liep 2 serving saline areas to produce rice-shrimp systems; The results of these studies are also valuable references for scientists on conservation of local rice genetic resources in the Mekong Delta, selection of plant varieties and other related fields
Trang 6CHAPTER 3: MEANS AND METHODS
3.1 Experimental materials
3.1.1 Place of research: some laboratories of Can Tho University For
the rice-shrimp system, the experimental field must have been cultivated according to the rice-shrimp system many years ago, with shrimp farming done in January–July and rice farming with shrimp farming from about August to December every year
3.1.2 Research materials included sources of local rice varieties from
the Can Tho University Gene Bank: Doc phung, Bo liep 2, Trang bo cau, Mong chim roi 3, Ba bui lun, Tet ran, Nang thom, Thom man, Nang quot bien 1, Nang co do 2, Tra long 2, Ba bui 2, Lun man, Nam tai 1, Mot bui 5, Nang cum 1, Thom lun mua, IR28 (saline standard), and Pokkali (salinity resistance standard)
Rice varieties included TN1 (Taichung Native 1 [BPH infection standard]) and Ptb33 (BPH resistance standard) received from the Southern Plant Protection Center
3.2 Research Methods
3.2.1 Collection and data for seed sources
Selection of locations and sample collection occurred in February 2014 Methods of seed collection included preparation of collection slips, sampling
of seeds at households, labeling, preliminary processing, and packaging
3.2.2 Cultivation, description, and evaluation of 41 rice varieties 3.2.2.1 Experiment 1: Evaluation of genetic characteristics
Location and time: Khanh Lam commune, U Minh district, land for two rice crops/year Time: June 2014 to June 2015
Methods: Experiments were arranged in a sequential manner without repetition The goal was to limit soil fluctuations affecting rice yield, so 41 local rice varieties (24 rice varieties from farmers and 17 rice varieties received from the Can Tho University gene bank) were coded using the random method (Table 3.2)
3.2.2.2 Characterization on morphology and molecular markers
Describe some morphological characteristics (IRRI, 2013)
Trang 7Table 3.2: List of 41 local rice varieties of experiment 1
a Evaluation of genetic diversity based on agronomic traits
Shannon index
Using the biodiversity index H’ (Shannon and Weiner's Index) according to the method of Shannon and Weiner (1963), the H index is calculated based on biomass (W) rather than the number of individuals (Ni)
H’ = Biodiversity Index (Shannon-Wiener index)
Wi = Biomass of species ith
W = Total biomass of all species collected in the field
Table 3.3: Criterion H' assessed (Duong Tri Dung, 2001)
Trang 83.2.2.3 Evaluation of salinity tolerance level of local rice varieties
Experiment 2 (Evaluation of salt tolerance of 41 rice varieties) was conducted at the Mekong Delta Development Research Institute using Yoshida nutrient solution Purification took place according to IRRI's saline purification procedure (1997) Time: Februrary through March 2015
3.2.3 Using SSR molecular markers to link salt tolerance (RM3412, RM493), BPH resistance (B121 and RM5479)
Experiment 3 (Identification of salt tolerance, resistance to brown planthopper) was conducted at the molecular biology laboratory of the Institute of Biotechnology, Can Tho University, from September 2015 to May 2016 Method: DNA extraction according to CTAB procedure (Rogers & Bendich, 1988) PCR reaction was by the PCR method using McCouch's SSR primer by agarose gel electrophoresis (1–3%)
3.3 Evaluation of some agronomic characteristics, yield components, and yield of rice varieties in the rice-shrimp system
Experiments 4 and 5 occurred in two districts of Thoi Binh and Tran Van Thoi, Ca Mau province, from July 2015 to June 2016 and involved the rice-shrimp system Method: completely random block with three iterations
To measure pH, EC, and field water, a plastic pipe with holes drilled for water
to seep through was placed in water at a depth of 0–20 and 20–40 cm in the soil (Figure 3.2) The experiements evaluated yield components and yield (IRRI, 2013); milling rate, length, and width of grain (mm) and rice classification according to IRRI (2013); and analysis of amylose according
to the method of Cagampang and Rodriguez (1980)
Figure 3.2: Water at 0–20 and 20–40 cm depth in the soil
Trang 93.4 Pure filtration of two varieties of rice to improve yield and quality
Experiments 6 and 7 took place in Khanh Lam, U Minh district, Ca Mau province, at a rate of two rice crops per year The experiments were conducted from July 2016 to July 2017 Experiment 6 was performed to compare the height, yield composition, yield, and grain quality of the Ba bong man line/variety rice Experiment 7 was performed to compare the height, yield composition, yield, and grain quality of Bo Liep 2 line/variety rice Methods: Experiments 6 and 7 were arranged in a sequential manner, without repetition
3.5 Narrow-area evaluation of two purebred varieties in the shrimp system in Ca Mau province
rice-Location: experiment 8-13 at U Minh, Thoi Binh anh Tran Van
Thoi district Time: from July 2017 to June 2018, in the rice-shrimp system Method: completely random block with three repetitions
3.6 Wide-area evaluation of two pure filtered varieties
Location: Experiment 14-21 at Kien Giang, Ca Mau, Bac Lieu and
Soc Trang province Time: From July 2018 to March 2019, in the shrimp model Methods: Experiments at each site were arranged in a
rice-completely randomized block design with three replications
3.7 Data processing methods
The field data were collected according to the standard system of rice genetic resources assessment of IRRI (2013) The Jaccard genetic similarity coefficient and UPGMA method in NTSYSpc 2.1 software were used for analysis In the analysis process, Excel, SPSS, and Past3 software were used to analyze the data
CHAPTER 4: RESULTS – DISCUSSION
4.1 Collecting local rice varieties
As a result, 24 rice varieties (Lun phen, Lun do, Lun phen hat nho, Mot bui lun 2, Lun vang, Lun phet, Lun hen, Lun sua, Lun can do, Lun can trang, Lun cao san do, Lun cao san trang, Trang phieu, Tep hanh, Mot bui lun, Mot bui do lun CM, Mot bui trang, Tai nguyen ST, Tai nguyen
CM, Nang quot bien, Soi lun, Ngoc nu, Ba bong man, and Mot bui do cao
Trang 10CM) were collected, cleaned, dried, cleansed of diseased seeds, and packaged for the documentation of genetic resources
4.2 Selection of local rice varieties for rice-shrimp farming
4.2.1 Evaluation of some morphological characteristics
Rice leaf morphology: The results show that the leaf blade
pubescence had three levels of expression: one variety had the glabrous (Tet ran), 39 varieties had the intermediate, and one variety had the pubescent (Nang thom) Leaf blade color had four expression levels: two varieties with light green, 32 varieties with green, six varieties with dark green, and one variety with purple tips Basal leaf sheath color had two levels of expression: 39 varieties showed green and two varieties showed purple lines (Ba bui 2, Tet ran) The leaf angle is just below the leaves with
40 rice varieties with erect angles and one variety (Nam tai 1) with horizontal The flag leaf angle had four expression levels: 19 varieties had erect, 14 varieties had intermediate, eight varieties had horizontal, and zero cultivars had descending Collar colors were light green (38 varieties), green (two varieties), and purple (one variety) For auricle color, 40 varieties showed as white and one variety showed as purple (Lun hen)
Morphology of the ligule: For ligule color, 41 varieties had white
(100%) For ligule shape, 39 of 41 varieties (95%) had truncate and and two varieties had acute to acuminate
Morphology of the culm (stem): The culm angle was erect (<300)
in 39 varieties and intermediate (~450) in two varieties (Nang quoi bien and Nang cum 1) The culm internode color was light gold in 38 varieties and green in three varieties (Nang thom, Bo liep 2, and Ba bui 2), and there were no varieties with purple lines The culm strength was expressed at all five levels: 11 varieties were strong (no bending), 21 varieties were moderately strong (most plants bending), four varieties were intermediate (most plants moderately bending), and five varieties were weak (most plants nearly flat)
Morphology of the panicle: The panicle type of rice varieties had
three forms: compact for three varieties (Ba bong man, Tep hanh, Thom
Trang 11man), intermediate for 37 varieties, and open for one variety (Nang thom) For the secondary branching of panicles, there were two levels: light with
24 varieties (59%) and heavy with 17 varieties For panicle exsertion of male sterile lines, 16 varieties had 0% extent of coverage of panicles by the flag leaf sheath, two varieties had 1–10% extent of coverage of panicles by the flag leaf sheath, 23 varieties had 11–25% extent of coverage of panicles by the flag leaf sheath, and no varieties had extent
of coverage of 26–40% or above 40% All the local rice varieties had the panicle axis showing as droopy (100%), with none showing as straight
Conclusion: Some morphological characteristics can be evaluated
with a high degree of diversity such as flag leaf angle, culm strength, and panicle exsertion of male sterile lines Some morphological characteristics can be assessed as non-genetically diverse, such as ligule color and panicle axis The others are classified as moderately diverse because each characteristic has two to three different levels of expression
4.2.2 Evaluation of some morphological characteristics
The quantitative characteristics of 41 local rice varieties were evaluated including culm length, leaf length, leaf width, ligule length, panicle/m2, seed/panicle, 1,000-grain weight, grain yield, white rice length, white rice width, chalkiness of endosperm, and amylose content Biodiversity index analysis results in all of these indicators They have a Shannon biodiversity index value of H' >3 According to Duong Tri Dung (2021), this shows that the varieties have good biodiversity According to Duong Tri Dung (2021), the H' index fluctuates from 1 to 6 He states that the H' index also depends on many other factors such as climate characteristics, latitude, relative altitude, and pollution level of the environment Tropical rainforests typically have very high H-index values of 5.06–5.40 compared with 1.16–3.40 for temperate forests and also for tropical plantations The H-index in heavily polluted water basins
is only 1 or less, while in clean water it may be 2–3 or higher
4.2.3 Grouping of genetic relationships of quantitative characteristics 4.2.3.1 Genetic diversity analysis of morphological characteristics
Trang 12Using principal component analysis (Figures 4.1a and 4.1b), we observed that the first axis mainly accounted for the variation in the germplasm (49.1%), followed by the second axis (21.9%)
This result indicated that plant height (group I) and panicles per square meter (group II) within the axes exhibit great influence on the phenotype of the population Moreover, the first five components accounted for 99.30% of the total variation, with components PCA3 (chalkiness of endosperm), PCA4 (leaf length), and PCA5 (amylose content) contributing 12.7%, 9.7%, and 5.90%, respectively In Figure 4.1c, there are 13 rice varieties in group I, including Mong chim den, Nang cum
1, Tet ran, Ba bui 2, Nang quot bien, Bo liep 2, Thom lun mua, Nang thom,
Figure 4.1 Cluster analysis of 41 rice germplasms (a) Proportion of information retained by each principal component (PCA1 Panicles per square meter; PCA2 Plant height; PCA3; PCA3 Chalkiness of endorsperm; PCA4 Leaf length; PCA5 Amylose content; PCA6 Grain per panicle; PCA7 Actual yield; PCA8 Grain weight) (b) Principal component analysis of 41 rice germplasm based on morphological traits (c) Heatmap analysis of the distribution of 11 quantitative morphological traits among 2 groups
Trang 13Doc Phung, Tai nguyen CL, Trang bo cau, Thom man, and Tai nguyen Group II has 28 rice varieties: Lun can do, Lun hen, Nam tai 1, Ngoc nu, Nang quot bien 1, Tep hanh, Mot bui do cao CM, Nang co do 2, Mot bui
5, Lun do, Lun cao san trang, Soi lun 1, Lun vang, Ba bong man, Lun phen, Lun phet, Mot bui lun, Trang phieu, Tra long 2, Mot bui do lun CM, Mot bui trang, Lun man, Ba bui lun, Mong chim roi 3, Lun phen hat nho, Lun sua, Lun cao san do, and Lun cao san trang In group I, Thom man had the lowest amylose content The heatmap obtained from cluster analysis (Figure 4.1c) shows that group I had 13 rice varieties which represented the plant height group, while group II tended to express mainly on panicles per square meter Thus, based on phenotypic traits, the analyses have been widely applied for studied genetic diversity (Liu et al., 2015; Veasey et al., 2008), and germplasm GenBank material can be identified (Islam et al., 2018) The findings from this study have potential benefits for the future use costal of Vietnam’s Mekong delta with a higher-yield rice molecular breeding program
4.2.3.2 SSR polymorphism among 41 local rice varieties
To obtain highly informative SSR markers, 50 SSR primers were tested in this study Following this, the PIC value was confirmed based on
a variation between 0.0 and 0.5 and above 0.5 for effective and informative markers In this study, PIC values absorbed from 50 SSR analysis mostly ranged from 0.00 to 0.49 The effective and informative primers were RM211, RM6329, RM127, RM253, RM24330, RM271, and RM286 The lowest PIC values were scored by 16 SSR primers (RM431, RM283, RM452, RM338, RM161, RM507, RM334, RM178, RM17749, RM162, RM455, RM125, RM11, RM536, RM144, and RM277)
According to the results as shown in Figure 4.5, the similarity coefficient given by Nei and Li (Nei, 1973) of relationships in this study ranged from 0.78 to 0.96 Additionally, 41 rice germplasms were officially divided into two main clusters The first cluster contained 39 germplasms (Doc phung, Lun can do, Lun can trang, Bo liep 2, Mot bui
do lun CM, Mot bui lun, Ba bong man, Lun cao san do, Lun cao san trang,
