Breeding for quality traits (protein,oil)
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This document discusses breeding crops for improved quality traits like protein and oil content. It covers topics like: - Quality traits can be morphological, organoleptic, nutritional, or biological. - Protein efficiency ratio and biological value are measures of protein quality in foods. - Breeding maize with higher lysine and tryptophan content led to the development of Quality Protein Maize varieties. - A case study describes using in vitro mutagenesis and selection with hydroxyproline to develop peanut varieties with over 55% oil content in kernels. - Breeding objectives for sunflower include seed yield, oil content, and modifying oil quality traits like fatty acid composition.
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Breeding for quality traits (protein,oil)
- 1. BREEDING FOR QUALITY TRAITS (PROTEIN, OIL) Presented by S.S. UDAYA KARTHIKA II. M. Sc.(Agri)-Genetics and Plant Breeding
- 2. QUALITY TRAIT MORPHOLOGICAL ORGANOLEPTIC NUTRITIONAL BIOLOGICAL SIZE COLOUR PALATABILITY TASTE AROMA SMELL JUICINESS PROTEIN OIL MINERAL VITAMIN PROTEIN EFFICIENCY RATIO BIOLOGICAL VALUE
- 3. PROTEIN EFFICIENCY RATIO • Gain in gram in body weight of an animal on consumption of 1 gram of protein • In cereals, high lysine content is directly proportional to high PER value BIOLOGICAL VALUE • Nitrogen retained in the body of animal • Total protein nitrogen absorbed by it from diet
- 4. NUTRITIONAL QUALITY ❖ PROTEIN QUALITY (AMINO ACID BALANCE) ❖ PROTEIN CONTENT ❖ PROTEIN DIGESTIBILITY ❖ OIL CONTENT ❖ OIL QUALITY (FATTY ACID COMPOSITION) ❖ VITAMIN CONTENT ❖ MINERAL CONTENT ❖ ABSENCE OF ANTINUTRITIONAL FACTORS
- 5. PROTEIN ORGANIC MACROMOLECULES LONG CHAIN OF AMINOACIDS LINKED BY PEPTIDE BONDS JOINING THE CARBOYL GROUP (-COOH) GROUP OF ONE AMINO ACID WITH AMINO GROUP (-NH2) OF OTHER AMINO ACID
- 6. ➢ PROTEIN – 20 AMINOACIDS ➢ AMINOACID SEQUENCE IS DETERMINED BY ✓ BASE SEQUENCE OF GENE ENCODING THAT PROTEIN ✓ FUNCTIONAL PROPERTIES OF PROTEIN ➢ NUTRITIONAL PROPERTIES DETERMINED BY ✓ COMPOSITION OF AMINOACID ✓ NOT BY AMINOACID SEQUENCE
- 7. PROTEIN QUALITY 8 ESSENTIAL AMINOACIDS ISOLEUCINE( Ile ) LEUCINE( Leu ) LYSINE( Lys ) METHIONINE( Met) PHENYLALANINE( Phe ) THREONINE( Thr ) VALINE( Val ) TRYPTOPHAN (Trp)
- 8. WHEAT ATLAS 66 – SOURCE OF PROTEIN CONTENT GENES OBTAINED GENES FROM “FRONDOSA” ▪ LEAF RUST RESISTANCE ▪ PROTEIN CONTENT LANCOTA – IMPROVING PROTEIN CONTENT ABSORB MORE SOIL NITROGEN ▪ HIGHER NITRATE REDUCTASE ACTIVITY ▪ THAN CHECK VARIETY “LANCER”
- 9. High lysine mutant Hiproly barley High lysine trait P721 Opaque mutant Opaque 2 gene Vivek QPM9
- 10. CASE STUDY 1
- 11. MAIZE-3rd major cereal crop Mostly consumed in Africa, Latin America, Asia ❖ Zein – most abundant protein ❖ Highly unbalanced in composition ❖ Deficient in lysine and tryptophan ❖ Cannot be synthesized in body
- 12. ➢ Poor net protein utilization ➢ Low biological value ➢ Cause malnutrition and leads to Kwashiorkar ➢ Insufficient essential aminoacids leads to under nutrition in ✓ Infants ✓ Lactating mother ✓ Pregnant mother
- 13. QPM is an improved variety of maize which contains higher amount of lysine and tryptophan in endosperm than normal maize
- 14. QPM DISCOVERY 1920-Spontaneous mutant of maize 1964-Homozygous recessive o2 allele Soft and opaque grains – Opaque2 (o2) Reduce zein fraction(50%) Lysine and Tryptophan proportion is doubled
- 15. ZEIN MAJOR GROUPS MINOR GROUPS 19 Kd & 22 Kd α zeins β-15 kD, γ-16 kD &27 kD, σ-10kD zeins
- 16. Opaque 2 gene Short arm of chromosome 7 Encodes leucine zipper(bZIP) class transcriptional factor Downregulate Cause opaque phenotype 22-kDa α zeins
- 18. PLEIOTROPIC EFFECTS OF THIS OPAQUE (O2) GENE Soft endosperm Damaged kernles Susceptibility to pest and diseases Reduced yield Inferior food processing
- 19. Due to its pleotropic effect ,opaque appearance and unpleasant taste which results in low grain yield
- 20. 3 Genetic systems involved in QPM breeding Recessive mutant allele of opaque -2 gene Endosperm hardness modifier gene Amino acid modifiers/gene
- 21. In 1976, Bjarnason et al, found set of QTLs in opaque-2(chromosome 7) to improve hard & vitreous kernels In 1980, Vasal et al, combined opaque-2 allele with QTLs to produce elite germplasm with hard kernel & increase in lysine & tryptophan content
- 22. Recently, In 2016, Liu et al.2016, found a 15.26 kb duplication segment (qy27) at 27 kDa γ zeins locus is reported to be a major modifier QTL to enhance the expression of protein & lead to endosperm hardness Multiple genes are involved in the control of aminoacid. Three loci are implicated in control of lysine level
- 23. CONVENTIONAL BREEDING (PHENOTYPIC SCREENING) MARKER ASSISTED SELECTION (GENOTYPIC SCREENING) Both genes o2 and endosperm modifier genes are selected DNA fingerprinting to help in matching physical properties of QPM. Light based screening 3 SSR markers –Foreground selection marker for o2 gene Seeds placed on Plexiglass surface above a light and screened endosperm DNA Extraction – identifying QPM plants early Human error or mis-classification may occur PCR based Co-dominant markers- robust, significant and reproducible Confirmation by molecular markers or amino acid content profiling CML 145*CML 212 Vivek QPM 9 BREEDING APPROACHES
- 24. MOLECULAR APPROACHES TECHNIC PLANT MATERIAL AIM REFERENCE SSR QPM & non-QPM Marker assisted introgression of o2 trait Manna, R;Okello, D.K. ISSR Primers and variety diagnostic markers QPM & non-QPM Molecular breeding program development Nkongolo,K.K;Daniel,G. SDS-PAGE QPM grain Protein identification Zhang,Z;Song,L. Proteomic analysis QPM inbred lines Pfunde,C.N. ISSR and RAPD markers QPM & non-QPM Assess the level of genetic variation Mbuya,K. SNP markers QPM lines Genetic diversity studies Pfunde,C.N. SNP-based genetic distance QPM inbred lines Heterotic grouping Badu-Apraku,B.
- 25. COUNTRY VARIETY DESIGNATION VARIETY TYPE AREA OF ADAPTION YEAR OF RELEASE India Shakthi OPV Tropical rainfed 1971 India Rattan OPV Tropical rainfed 1971 India Protina OPV Tropical rainfed 1971 India Shaktiman 1 Hybrid Tropical rainfed 2001 India HQPM1 Hybrid Tropical rainfed 2006 India HQPM7 Hybrid Tropical rainfed 2008 India Vivek QPM9 Hybrid Mid- Altitude/Highland, rainfed 2008 List of QPM varieties released till 2015
- 26. OILS AND FATS Esters of glycerol & aliphatic straight chain fatty acids CH2OH R1-COOH CH2OOC-R1 | | CHOH + R2-COOH CHOOC-R2 + 3H20 | | CH2OH R3-COOH CH2OOC-R3 (Glycerol) (Fatty acids) Triglyceride(natural oil/fats) Esterification
- 27. Types and proportion of fatty acids present in the triglycerides Saturated fatty acid crops Oil palm Coconut Cocoa Oleic-Linoleic Acid Crops Maize Groundnut Soybean Safflower Sesame Cotton Erucic Acid Crops Rapeseed Turnip rape, Toria Mustard
- 28. Breeding Approaches • Domestication • Selection • Mutation • Hybridization • Interspecific Hybridization Biotechnological Approaches • Micropropagation • Somo clonal Variation • Anther culture • Somatic hybridization • Genetic engineering
- 29. MUTATION BREEDING IN OILSEED CROPS Oilseed crops – economically important crops Major crops- Soybean, rapeseed, Groundnut, Cottonseed Minor crops- Linseed, Fenugreek
- 30. Three step process of mutation breeding a)Inducing mutations b)Screening for putative mutant candidates c)Mutant testing and official release Mutation based plant breeding aimed to upgrade well adapted varieties by altering one or two major traits Mutation breeding-most viable method. Induced mutation / molecular mutation breeding-improving food security
- 32. VARIETIES YEAR CROP IMPROVED CHARACTER TG1(Vikram) 1973 Groundnut Oil content and TMV resistance RLM 198 1975 Mustard Oil content, Early maturity RLM 514 1980 Oriental mustard Oil content, Less erucic acid RL 1359 1987 Chinese mustard Oil content, Short duration TGS 1(Somnath) 1989 Groundnut Oil content , Sequential flowering UMA 1990 Sesame Early and uniform maturity, Oil content OIL CROP VARIETIES IN INDIA
- 33. CASE STUDY 2 The use of invitro mutagenesis and directional OP based selection to develop peanut varieties with increased oil content in kernels
- 34. Materials and Methods 1.Peanut tissue culture: a)Basal medium b)Embryo induction medium c)Mutagenesis medium-mutagen(pingyangmycin) d)Embryo germination & screening medium *6-benzyl amino purine(BAP) *Hydroxyproline(HYP)-OP regulator All the media are adjusted to pH 5.8 and autoclaved at 120 C for 20 mins.
- 35. 2.Determination of optimal conc of the mutagen Cotyledons are removed Embryos are surface disinfected and soaked in sterile water for 10-12 hour. Explants dissected from embryos and cultured in culture bottle (36 explants per bottle) containing 45mL of mutagenesis medium supplemented with PYM at 0,1,2,3,4 or 5 mg/L. 4 weeks later somatic embryo induction rates were calculated by formula: Somatic embryo no of explants forming somatic embryos *100 Induction (%)= No of explants The embryo induction rate was appx 50% with no PYM, so we selected 4mg/L PYM as the optimal conc for mutagenesis
- 36. 3.Mutagenesis, selection, grafting, and transplantation of regenerated plants with HYP tolerance After regeneration of plantlets from somatic embroys, they are subcultured. 1.5 cm height regenerated plant are grafted to the rootstock from Huayu 20 After 3 days sterile culture, they are transferred to pots containing mixture of soil: vermiculite :turfy soil(1:1:1) Three weeks later, they are transplanted in the field and seeds of M1 generation is harvested.
- 37. 4.Selection of mutants from progeny of HYP Tolerant plants Individual plants are selected from M2 to M5 generations. Seeds from the generations and parents are planted in the field. Selection was performed based on plant type, branch number, pod shape, and pod number per plant. 5.Measurement of oil content Mature dry seeds were sent to the Quality Inspection and Test Centre for Oilseeds and measured using residue method.
- 38. 6.Yield comparison test The offspring of M6 generation with higher yield and oil content greater than 55% was chosen for the yield comparison test.M7 and M8 generations accessed in separate experiment using RCBD with 3 replications .Huayu 20 was used as a control 7.Variety regional test for variety registration Tests were performed using RCBD with 3 replications at 6 locations and Baisha 1016 used as control. At maturity pods are harvested and weighed. 8.Statistical analysis
- 39. ➢ Here, we used HYP to increase the osmotic pressure from 77.0 to 85.5 mmol kg–1 in the embryo germination and screening media. ➢ Water generally flows from low OP to high OP regions. ➢ Because of the increased OP in the media, we assumed that the somatic embryos or regenerated plantlets with normal oil content would die due to water loss, and only those with dehydration tolerance would be able to survive. ➢ When the somatic embryos that survived in vitro mutagenesis were incubated and screened on somatic embryo germination medium supplemented with 6 mmol L–1 HYP (OP=85.5 mmol kg–1), only about 0.5% survived, and 15 regenerated plantlets were obtained. RESULTS
- 41. Based on the results for yield and oil content presented here, three lines were submitted for regional testing. Line 4-11-1-1 was tested in Anhui in 2015; lines 1-3-6-3 and 2-1-9-1 were tested in Liaoning in 2016 and 2017, respectively. In each of these provinces, tests were conducted at six locations with Baisha 1016 being included as a control. The average yield was 16.6, 13.9 and 14.4% higher for lines 4-11-1-1, 1-3-6-3 and 2-1-9-1, respectively, than for the control variety.
- 42. These three lines were named varieties: Yuhua 4 (line 4-11-1-1), Yuhua 9 (line 1-3-6-3) and Yuhua 14 (line 2-1-9-1). The oil content was 57.9, 61.1 and 59.3% for Yuhua 4, Yuhua 9 and Yuhua 14, respectively. Among all peanut varieties currently planted worldwide, Yuhua 9 has the highest oil content. Conclusion This study demonstrated that the combination of PYM as a mutagen and in vitro screening with HYP is a promising way to create new peanut mutant and obtain varieties with high oil content. It provides an effective approach for peanut breeding and will be useful for breeding peanut varieties with higher oil content.
- 43. Photographs of representative somatic embryos that formed from embryonic leaflets on the induction medium without (A) or with (B) pingyangmycin (PYM) at 4 mg L–1. Photographs of representative somatic embryos and plantlets that survived pingyangmycin (PYM) mutagenesis and subsequently survived on a medium supplemented with 6 mmol L–1 hydroxyproline (HYP) after 3 wk (A), 5 wk (B) and 8 wk (C)
- 44. Photographs of representative grafted plants with hydroxyproline (HYP) tolerance and their appearance when transplanted in the field and at pod harvest. A and B, grafted plants with HYP tolerance. C, No. 14 regenerated plant had purple stems and branches. D, an HYP-tolerant plant and its pods (the arrow points to the graft union)
- 45. Photographs showing representative trait variation in the M2 generation. A, Huayu 20. B, progeny of the No. 14 plant. C, progeny of the No. 8 plant. D, pods from the No. 7 plant.
- 46. Perspectives of Breeding for altering Sunflower Oil quality to obtain novel oils Sunflower competes Soybean, Groundnut, Rapeseed Good quality of oil – PUFA – reduce cardiac risk Oil content – 35-45%
- 47. Concept of Oil quality is changed by ➢ Saturated fatty acid composition ➢ Triacylglycerol profile ➢ Tocopherol content & composition OIL QUALITY “Palmitic acid” – undesirable- detrimental atherogenic effect- rises serum cholesterol
- 48. High oleic acid % High linoleic acid Low linoleic acid Linoleic acid- essential for human High oleic hybrid –above 70% 1.Seed yield/unit area 2.Oil content 3.Oil quality BREEDING OBJECTIVE
- 49. Effect of Environmental factors Instability of genotypes across environment may be ❖ Planting location ❖ Planting date result in different during maturation stage ❖ High or low rainfall in growing season ❖ Presence of modifier genes Environment variability in oil content Genotypic factors variability in fatty acid composition
- 50. Early sowing - Oleic acid Linoleic acid At seedling stage, High night temperature Oleic acid Linoleic acid Low rainfall Accumulation of oil in seed requires a)below 25 C mean daily temperature b)sufficient soil moisture c)absence of disease Use of wild species: Sunflower lines with best oil concentration and quality improvement Helianthus deserticola (desert sunflower) Helianthus anomalus (sand sunflower)
- 51. Effect of mutation 1st artificial mutation by Soldatov (1976) 2.5% Dimethyl sulphate VNIMK Pervenets High oleic variety X-rays Sunflower seeds 30% palmitoleic acid Sodium azide Sunflower seeds 35% stearic acid
- 52. Genetic basis of sunflower ❖ P1, P2, P3 – 3 genes in control of high palmitic acid ❖ Es1, Es2, Es3 – genes in control of stearic acid ❖ O11 – principal gene – control of high oleic acid content ❖ Tph1, Tph2 – control altered tocopherol composition ❖ Two potential QTLs for OA & LA content identified with markers ORS 762 & HO-Fsp-b explained more than 57.66-66.6% of phenotypic variation.
- 53. These markers /QTLs useful in MAS breeding programe to improve oil quality Molecular markers for some of the traits have been developed for High stearic & oleic acid content High beta & gamma tocopherol contents Polymorphic markers applied for selection in backcross b/w high oleic donor & 2 low oleic recurrent parents.
- 54. CONCLUSION ➢ Quality protein maize has nutritional quality can alleviate protein malnutrition in maize dependent countries. ➢ Though conventional methods of breeding are still in use ,marker assisted breeding with markers for foreground selection can expedite good breeding process in all crops. ➢ This saves time, money and labour resources with high genetic gain.