Dr. Mirza Mofazzal IslamPrincipal Scientific Officer

Plant Breeding DivisionBangladesh Institute of Nuclear Agriculture (BINA)

Mymensingh, BANGLADESH

Development of Salt Tolerant Rice Variety(s) through Induced Mutation and Marker

Technology at BINA

07 July 2010

Bangladesh Institute of Nuclear Agriculture (BINA)

Mymensingh

WELCOME

Salinity Problem

World wide, salt affected areas = 400 – 950 m. ha (Lin et al., 1998).

8.5 x 106 haUSA

86 x 106 haArgentina

21 x 106 haParaguay9 x 106

haChile

357 x 106 haAustralia

37 China

24India

11Pakist

an

11 x 106haEthiopia

13 x 106haIndonesia

170 x 106 haRussia In Asia, only rice lands =

54 m. ha

Another 9.5 m. ha of saline soil => managed by irrigation drainage and by chemical treatment….i.e., too costly.

Need salt tolerance varieties.

Symptoms…….

Stunted growth.

Reduced number of tillers.

Reduced root growth.

Rolling and dying of leaves, leaf tips.

Decrease Photosynthesis.

Sterile spikelets (delayed flowering)

Result to ………..

Yield Reduction

BANGLADESH

Noakhali

Satkhira

Saline areas

Satkhira (Fellow lands of Coastal saline area) Bangladesh

Satkhira (Coastal saline area) Bangladesh

Satkhira (Coastal saline area) Bangladesh

To develop salt tolerant rice varieties through induced mutation and marker-assisted selection

To enhance rice production for food security and improve

livelihood of farm community

Objectives

End Users

Farmers, consumers and particularly people in

saline prone areas will be benefitted by adopting

salt tolerant rice variety(s)

Past and Present Country Efforts to address the need

In the past, Bangladesh Government allocated money for

National Agricultural Research Institutes (NARS) and national

universities for pursuing institutional core research to develop

salt tolerant crop varieties

Besides government efforts, International Rice Research

Institute (IRRI) and other international donors provided funds

to pursue research on development of stress (salinity, drought

and submergence) tolerant crop varieties

Bangladesh Government gives more emphasis on global warming and food security.

Considering the significance of the salinity problem in the country, Bangladesh Government recently initiated several research programmes towards developing high yielding salt tolerant rice varieties that can be grown in the salt affected areas.

Development projects such as “Strengthening research for improved high yielding climate resilient rice varieties”, “Agricultural Research and Development programme for Sustainable Food Security” funded by World Bank through the Global Agriculture and Food Security Program (GAFSP);

“Agricultural Research: Vision-2030 and beyond” funded by Bangladesh Agricultural Research Council (BARC) recently approved by the Government of Bangladesh.

At present, due to climate change gradually inlands are becoming more saline and the salinity level frequently reached up to 30 dS/m (extremely saline).

Recently we developed one mutant rice variety 'Binadhan-7' as early maturing, fine grain and high yielding.

We want to introgress salt tolerant gene in this variety through marker-assisted backcrossing.

We need technical and financial supports to do such

research works.

At our institute, we have also on-going institutional core funded project (with limited budget) for pursuing research on 'Development of salt tolerant rice varieties'.

We need to develop highly salt tolerant rice varieties with desirable yield. Despite the weak economic context, Bangladesh Government has taken many policies for the improvement of agriculture sector.

But the total budget for agriculture sector in our country is not sufficient for advanced research yet.

Role of Nuclear Technology

Irradiation will be applied on popular rice varieties and landraces to create variability for getting salt tolerant mutants.

Selected mutants will be used as donor/recipient parent in marker-assisted selection.

Irradiation technique will be complement with marker-

assisted breeding.

Role of IAEA

The capacity of the institute in advanced techniques in mutation breeding and marker technology through training/fellowship of young scientists and upgrading of facilities is needed.

To acquire advanced knowledge and information in mutation breeding and biotechnology through scientific visits are also essential to boost up the institute’s current research programme.

Mutagenic chemicals, biotech chemicals, consumables and some essential biotech equipments are urgent needed

for institute’s biotech lab.

Procurement system of mutagenic and biotech chemicals/consumables is very complex because of some bureaucratic policy and time consuming procedures.

It is expected that IAEA would provide support in the procurement of equipments, chemicals/consumables, training of manpower in advance laboratories and involve of

experts/consultants to develop research programmes.

Partnership

International Rice Research Institute (IRRI) is collaborating with our institute to some extent for developing stress (salinity, drought and submergence) tolerant and HYVs of rice.

It is expected that IRRI would provide rice germplasm,

technical expertise and training.

Physical Infrastructure and Human Resources

We have office buildings, experimental natural fields (on-stations and on-farms), glasshouse, biotechnology laboratory and cobalt-60 source.

It is expected that four plant breeders (senior to junior level), three Para Scientists/Assistant Scientists, three Field/ Lab Attendant and several Field Laborers of Plant Breeding Division of our institute will be involved in the project.

Mandate/Functions of the Institute

Undertake research adopting nuclear techniques for the purpose of ensuring a stable and productive agriculture through evolution of new varieties of crops

Scientific management of land and water

Development of appropriate technology to improve quality and quantity of crops and

Development of methods for control of disease and insect and management of pests

Undertake agronomic and soil-plant studies

Carry out demonstration tests or trial-runs of new varieties of crops and their management practices

Publish annual reports relating to activities of the Institute

Publish agriculture manuals, monograms, bulletins and other literature relation to crop research

Train research and extension officers in the improvement technology of crop production

Provide post-graduate research facilities

Organize seminars, symposia and workshops on selected problems relating to agriculture and agricultural research and technology

Undertake research programmes in collaboration with other national and international agencies and organizations

Enter into bilateral agreements with foreign institutions for academic degrees, specialized training, scientific visit and exchange programmes and

Perform such other functions as may be necessary for the purposes of this Ordinance

Sl.No.

Programme Area Implementing Divisions

1. Crop Improvement through Induced Mutation

Plant Breeding

2. Biotechnology Plant Breeding, Plant Pathology, Entomology, Soil Science and Crop Physiology

3. Soil Management and Biofertilizer

Soil Science

4. Irrigation and Water management

Agricultural Engineering

5. Pest Management Entomology, Plant Pathology

6. Physiological Aspects of Crop Productivity

Crop Physiology

7. Crop Management and On-farm research

Agronomy

8. Technology Transfer and Impact Assessment

Training, Communication and Publication

Program Area

Name of CropName of the variety

Season Main characteristics

Yield potential

(t/ha)

Av. yield (t/ha)

RICE Iratom-24 Boro and aus

Early, high yield, long grain 7.0 6.5

Binasail T. aman Tall, moderate early, moderate yield, low input requirement and suitable for late planting after flood

5.5 4.2

Binadhan-4 T. aman Early maturing, tall with long and slender grain 6.0 4.7

Binadhan-5 Boro High yield 7.0 6.0

Binadhan-6 Boro High yield 8.0 6.5

Binadhan-7 Aman Early, high yield 5.5 4.5

JUTE Atompat-38 Kharif-I High fiber yield with prominent genetic marker. 3.0 2.8

Binadeshipat-2 Kharif-II High fiber yield. 3.5 3.0

Binapatshak-1 Kharif-I Short and leafy plants used as vegetables (shak) 4.0 3.5

CHICKPEA Hyprosola Rabi High yield, high protein content. 1.5 1.4

Binasola-2 Rabi High yield, bold seeded. 1.7 1.5

Binasola-3 Rabi Early, tolerant to Botrytis grey mould and root rot. 2.3 1.6

Binasola-4 Rabi High yield, tolerant to Botrytis grey mould and root rot, bright seed coat

2.5 1.7

Achievements

Nameof crop

Name of the variety

Season Main characteristics Yield potential

(t/ha)

Av. yield (t/ha)

MUNG-BEAN

Binamoog-1 Winter High yield, tolerant to Cercospora leaf spot and other diseases

1.0 0.9

Binamoog-2 Summer High yield, tolerant to Cercospora leaf spot and Yellow Mosaic Virus (MYMV)

1.5 1.1

Binamoog-3 Winter High yield, tolerant to CLS and MYMV 1.1 1.0

Binamoog-4 Winter High yield, early, dwarf, tolerant to MYMV 1.2 1.1

Binamoog-5 Summer High yield, tolerant to MYMV 1.9 1.5

Binamoog-6 Summer Early 1.5 1.4

Binamoog-7 Summer High yield 2.0 1.8

GRASS-PEA

Binakhesari-1 Rabi High yield with low BOAA. 2.4 1.9

LENTIL Binamasur-1 Rabi High yield, rust resistant; black seed coat. 2.0 1.8

Binamasur-2 Rabi High yield 2.1 1.9

Binamasur-3 Rabi High yield 1.9 1.8

BLAC-GRAM

Binamash-1 Rabi High yield, tolerant to YMV and CLS 1.2 1.0

MUST-ARD

Safal Rabi High yielding & tolerant to Altenaria disease 2.4 1.70

Agrani Rabi Early, high yield, tolerant to Altenaria disease 2.60 1.75

Binasarisa-3 Rabi Dwarf, early, high yield 2.4 1.8

Binasarisa-4 Rabi Dwarf, early, high yield 2.5 1.9

Binasarisa-5 Rabi High yield, tolerant to salinity 2.1 1.6

Binasarisa-6 Rabi High yield, tolerant to salinity 2.2 1.5

Name of Crop

Name of the variety

Season Main characteristics Yield potential

(t/ha)

Av. yield (t/ha)

SESAME Binatil-1 Kharif-I

Uniculm plants with cream colour seed coat, high yield

1.4 1.0

GROUNDNUT Binachinabadam-1

Summer &

winter

Bold seeded, high yield 3.7 / 2.4

2.6 / 2.0

Binachinabadam-2

Summer &

winter

Short duration, high yield and resistant

to diseases

3.2 /1.7

2.5 /1.2

Binachinabadam-3

Summer &

winter

Short duration, bold seeded, high yield and resistant to diseases

3.0 / 1.6

2.4 /1.1

TOMATO Bahar Winter High yield, bigger fruit size, fleshy and tasty

75.0 65.0

Binatomato-2 Summer

Fruit shape round with small ridges. Can be grown without using hormone

42.0 38.0

Binatomato-3 Summer

Bigger fruit size, semi-round fruitwith prominent ridges and furrows.

Hormone is not required

45.0 40.0

Binatomato-4 Winter Round and smooth fruit 80.0 77.0

Binatomato-5 Winter Early, long and smooth fruit 72.0 69.0

Progress in breeding for salinity tolerance has been slow due to:

Limited knowledge in the genetics of tolerance

Complex – tolerance mechanisms

Inadequate screening techniques Low selection efficiency

Poor understanding of salinity and environmental interactions

At present, progress in revealing the genetics and mechanism of salinity tolerance in rice (Gregorio and Senadhira, 1993; Lee, 1995).

Selecting efficiency for salinity tolerance under field conditions remains very low.

Salinity tolerance is governed by polygenes or QTL.

Classical method cannot detect single gene locus associated with quantitative traits, their location in chromosome or their relationships with other genes.

Low selection efficiency in conventional breeding approach…..

takes almost 20 years and >10years by rapid anther culture.

Marker assisted selection (MAS) can analyze quantitative traits and identify individual genes controlling the traits of interest.

MAS is faster, more efficient and cost effective.

Current Research Thrust

Making the research programme future demand oriented

Linked to national development objectives

Ensuring stable and sustainable production systems

Ensuring securities in food, nutrition, employment and income, and other felt needs of the farming community through downstream agro-industries and processing

Conserving environment and natural resources based against over-use and abuse

Strengthening R & D activities

Marker-assisted selection for salt tolerance using

BINA mutants/lines

Total 428 germplasm

Out of 428 germplasm, 15 were tolerant and 20 were

moderately tolerant

Bangladeshi landraces- 385HYV – 15BINA mutant- 25Tolerant checks- 3 (Pokkali, Nonabokra and Horkuch)

Screening at seedling stage

SALINIZED SETUP

NON-SALINIZED SETUP

SCORE OBSERVATION TOLERANCE

1 Normal growth, no leaf symptom Highly tolerant

3 Nearly normal growth, but leaf tips or few leaves whitish and rolled

Tolerant

5 Growth severely retarded; most leaves rolled; only few elongating

Moderately tolerant

7 Complete cessation of growth; most leaves dry; some plants dying

Susceptible

9 Almost all plants dead or dying Highly susceptible

Modified standard evaluation score (SES)

Results of Seedlings Stage

► Salt tolerant : 5 lines

► Moderately tolerant : 17 lines

Based on SES scoring for salt tolerance

Screening at reproductive stage atBINA glasshouse

Screening at reproductive stage atBINA glasshouse

Several crosses have been attempted to develop salt tolerant rice varieties

Cross-1: Binadhan-5/Horkuch

131 F4 rice lines were tested for salinity tolerance at the

seedling stage

Selected 38 F4 lines were tested for salinity tolerance at the

reproductive stage

Selected 38 F4 lines also were grown at Satkhira and BINA HQ 15 F6 lines were selected and grown in AYT

Cross-3: Binadhan-5/ Pokkali

22 F3 lines were tested for salt tolerance at the seedling stage

Cross-2: Mut-1-1 / Pokkali

54 F3 plants were selected and tested with molecular markers

35 F4 plants were tested for salinity tolerance at the seedling stage

22 F4 plants were selected and grown in PYT in F5

Cont’d

Proposed Marker-Assisted Strategy Backcrossing program at BINA

Binadhan-7 × FL 378

F1 × Binadhan-7

BC1F1

BC1F2 × Binadhan-7

BC2F1

BC2F2 × Binadhan-7

BC3F1

BC3F2NIL

Marker-Assisted Backcrossing

Marker-assisted backcrossing were made between

Binadhan-5/FL-378= 105 BC1F1 populations

Binadhan-5/FL-478= 203 BC1F1 populations

New crosses Binadhan-7/FL-478 = 483 F1 seeds

Binadhan-7/FL-378 = 541 F1 seeds

Binadhan-5/FL-478 = 35 F1 seeds

Binadhan-5/FL-378 = 133 F1 seeds

Field trials of selected lines at Satkhira

Binadhan-5/Horkuch: 15 F6 lines

Mut-1-1/Pokkali: 22 F5 lines

Carbon Isotope Discrimination

Plants tend to selectively fix the light and most common stable isotope form of carbon (12C) and tend to exclude the heavy form (13C).

The isotopic ratio 13C to 12C in plant tissue is less than the isotopic ratio of 13C to 12C in atmospheric CO2.

This discrimination mainly occur during fixation by Rubisco and to lesser extent during diffusion through stomata since the heavy form move slower.

CID contd.

The discrimination is greater when stomata are widely open but tend to decrease when stomata partially close as under drought/salinity.

Reduction of stomata usually result in high WUE.

Selection for greater ∆ or lower ∆ - will be effective for improving salinity tolerance.

Something we need to test………

Flag leaf samples were collected before panicle initiation from both salinized and non-salinized setups.

Leaves were dried in oven at 700C for 3 days.

Dried leaves were ground in a mill to find powder.

C13 were analyzed at IRRI using ratio mass spectrometer.

Carbon isotope discrimination (Δ)

Under non-salinized condition heritability low

Number of filled grains/plant (14.6) 1000-seed weight (72.7) Grain yield/plant (29.9)

Based on Heritability (%) at the reproductive stageUnder salt stress heritability high

Number of filled grains/plant (58.1)

1000-seed weight (85.9)

Grain yield/plant (45.3)

y = -0.0505x + 21.342

R2 = 0.1675

20.4

20.6

20.8

21.0

21.2

21.4

0 2 4 6 8 10

SES Score

Δ13

C (%

)

Seedling Stage

Association between ∆L vs SES score was found as negative under salt stress at the seedling stage. These signify that highly salt tolerant lines are responsible for higher value of ∆L.

Reproductive stage

There was significant positive association between ∆F vs total dry matter, number of filled grains and grain yield/plant of 38 F3 lines under salinized condition at the reproductive stage

y = 0.0619x + 21.737

R2 = 0.0544

18.00

19.00

20.00

21.00

22.00

23.00

24.00

0 5 10 15 20

Total dry matter

Δ13

C (

%)

Association between ∆ 13C vs total dry matter

y = 0.0047x + 21.687

R2 = 0.0761

19.0019.5020.0020.5021.0021.5022.0022.5023.00

0 50 100 150 200

Number of filled grains/plant

Δ13

C (

%)

Association between ∆13C vs number of filled grains/plant

y = 0.1574x + 21.986

R2 = 0.0935

21.4021.6021.8022.0022.2022.4022.6022.8023.00

0 0.5 1 1.5 2 2.5 3 3.5

Grain yield/plant

Δ13

C (

%)

Association between ∆13C vs grain yield/plant

Future Challenges And Frontier Research

Crop Improvement

Development of saline, drought, submergence and temperature tolerant crop cultivars

Development of disease and insect pest tolerant cultivars

Genetic conservation and biodiversity

Development of hybrid and super varieties

Management of transgenic, hybrid and super varieties

Soll and Water Management: (Radio-isotopes like P32, Zn65, C14, N15, H3, O18 etc. have been in use at BINA)

Pest Management issues: (integrated use of biological and botanical pesticides, cultural and irradiation techniques)

Crop Management and On-farm Research: (Biological amelioration and agronomic practices for soil fertility versus sustainable productivity

Technology adoption and regeneration

Others:

Thrusts in long-term upstream research

Encouraging an integrated multi-disciplinary approach in research

Continuously updating and implementing scientific man-power development plant

Improving the terms and conditions of appointment of scientists for their productive utilization and increased output

Introducing better research management procedures for the improved efficiency and accountability.

Future challenges

Flood, Dhobaura, 2009

Flood, Dhobaura, 2009

Drought

More challenges

- Quality enhancement

- Multiple Tolerant/Resistant to biotic/abiotic

stresses

- New Plant Type (NPT) or Ideotype Breeding

- Allele or gene mining (GENE DISCOVERY)

Genotypic observations (MAS)

1Kb+ P1 P2 1 2 3 4 5 6 7 8 9 10 11 12

(b)

146-197 bp→

290-312 bp →

(a)

Fig. Banding profiles of 24 F2 population derived from the cross Binadhan-5 x Bawoi Jhak using primer (a) RM334 and (b) RM594; Lane 1: 1 kb+, Lane 2: Binadhan-5 (P1), Lane 3: Bawoi Jhak (P2), Lane 4-15: F2 lines

Thank you