ABSTRACT

The oil of low erucic acid Brassica juncea contains lower levels of C18:1 and higher levels of C18:2 fatty acids than does the commonly grown canola species, B. napus and B. rapa. Modified fatty acid breeding lines of B. juncea , having a fatty acid profile similar to that of B. napus, were selected. In this paper we report our studies on the inheritance of the modified fatty acid composition in B. juncea.

The unmodified material had a fatty acid composition of approximately 50% C18:1 and 30% C18:2, whereas the modified lines had approximately 60% C18:1 and 20% C18:2. F₁ seeds had a fatty acid composition similar to that of the unmodified parent, indicating that the modified composition is recessive to the normal composition, and that the fatty acid composition of the seed is under the genetic control of the developing embryo rather than the maternal plant, as is common in the Brassicas.

Individual F₂ seeds from self pollinated F₁ plants segregated into two slightly overlapping groups, showing a 3:1 ratio for normal versus modified fatty acid profile. In both the F₂ data set the C18:1 values were a little higher and the C18:2 values were a little lower than would be expected from a perfect monogenic dominance model, possibly suggesting some minor additive effects.

In a practical plant breeding program the high C18:1 - low C18:2 characteristic can be treated as a single gene, recessive to the "wild type" low C18:1 - high C18:2. The C18 fatty acid composition is under embryonic control, allowing the use of 1/2 seed analyses. Use of C18:2 concentration gives a better resolution of the two genotypes than does the use of C18:1 concentration.

KEYWORDS Oleic acid, Linoleic acid, Inheritance

INTRODUCTION

In order to be acceptable into the Canadian canola commodity market, Brassica juncea not only needs to meet the canola standards for erucic acid content and glucosinolates (Amendments to Seeds Regulations of the Canadian Seeds Act 1987), but also needs to have a fatty acid profile in the oil similar to that of the conventional canola species B. napus and B. rapa. The requirements for low erucic acid and low glucosinolate content have been met (Kirk and Oram 1981, Love et al 1990), but the fatty acid composition of most low erucic acid B. juncea lines differs significantly from that of the conventional canola species. In this paper we present data on the inheritance of the C18 fatty acid profile in B. juncea.

MATERIALS AND METHODS

Modified fatty acid (MFA) breeding lines of B. juncea, having a fatty acid profile similar to that of B. napus, were identified within the oilseed breeding program at the Saskatchewan Wheat Pool. Lines with a normal fatty acid profile (NFA) were from the breeding program at the Agriculture Canada Research Farm at Beaverlodge. All data presented here is from greenhouse grown material. Fatty acid analyses were by gas chromatography of the fatty acid methyl esters. The NFA material had a fatty acid composition of approximately 50% C18:1 and 30% C18:2, whereas the MFA lines had approximately 60% C18:1 and 20% C18:2.

Experiment 1

Initial cross Six crosses were made, using four different MFA female parent plants and six different NFA male parent plants. In addition the parental plants were self-pollinated. Fatty acid analysis was conducted on between 6 and 19 single seeds of each cross as available and 20 seeds of each self-pollinated parent plant. Average values are reported in Table 1. With the numbers of analyses available it was also possible to examine the variation from seed to seed in individual genotypes.

F₁ grow-out From each F₁ cross and each selfed parent (S₁) , four plants were grown and self pollinated. Typically twenty single F₂ seeds from each F₁ plant were analysed for fatty acid composition, as were 5 seeds from each of the parent S₁ plants¹.

F₂ grow-out A population of F₂ plants (16 plants from each of 6 selfed F₁ plants) was then grown, along with plants from each selfed parent (S₁) (16 plants from each of 10 parents). All individual plants were self pollinated. The harvested seed was analysed on a single seed basis, 20 seeds per F₂ plant and 5 seeds per S₁ parent plant, where available.

Experiment 2

To confirm the embryonic control of FA profile, the original cross was repeated in a second experiment, using 10 NFA and 14 MFA plants. Reciprocal crosses were made, and also self pollinated seed was produced. F₁ and selfed parental seeds were analysed for FA profile on a single seed basis (averages reported in Table 2).

RESULTS AND DISCUSSION

In total the parental material was grown and analysed four times. Very little difference in composition was detected between each grow out, and the observed distributions for the parents grown with the F₂ grow out are presented as typical (Figures 1-3). The parents were quite well resolved as far as C18:1 and C18:2 percentage, but there was considerable overlap in the C18:3 percentages.

In experiment 1 the F₁ seeds resembled the NFA parent for C18:1 and C18:2 percentages on an overall average basis (Table 1). Some individual seeds however had C18:1 or C18:2 contents which resembled the MFA parent, as a result the F₁ distribution exhibited a tail/smaller peak in the direction of the MFA composition (Figures 4 & 5). The C18:3 distributions of the parents and F₁ were not greatly different.

The F₂ seeds showed a bimodal distribution for C18:1 and C18:2 percentages, with a possible 3:1 ratio of NFA:MFA type (Figs 6 & 7). Again there was no distinct variation in C18:3 concentration.

The fatty acid distribution of the S₁ plants (Figures 1-3) was used as a basis for categorizing the F₂ plants. The C18:2 distribution was the best for separating the two classes. Each F₂ plant was categorised as MFA, NFA, or segregating based on the 20 single seed analyses, using the presence of C18:2 less than 20% to indicate a MFA seed, and greater than 30% to indicate a NFA seed. Using these criteria the F₂ population was estimated at 9 MFA, 48 segregating, and 34 NFA. There is a clear deficiency of MFA types for a 1:2:1 ratio which would be expected from the single gene situation which is suggested by the single F₂ seed analyses, with a significant χ² value (P<0.05). In the field poor vigour has been observed in breeding lines from these crosses which combine the MFA profile and low glucosinolates. Possibly germination failures or accidental selection against the MFA types may have resulted in this MFA type deficiency (2 seeds were planted per pot, and thinned to one after germination).

In experiment 2 the fatty acid profiles of the individual seeds were quite similar to the NFA parent (Table 2), although there did appear to be some slight skewing of the profiles towards the male parent composition for C18:1 and C18:2 (Figures 8 & 9), and towards the female parent composition for C18:3 (not shown).

Conclusion

In a practical plant breeding program the high C18:1 - low C18:2 characteristic can probably be treated as a single gene, recessive to the “wild type” low C18:1 - high C18:2. The C18 fatty acid composition is under embryonic control, so generation time may be saved if required by the use of 1/2 seed analyses. The C18:2 concentration gives a better resolution of the two genotypes than does the C18:1 concentration. The low C18:1 - high C18:2 genotypes seem to be slightly variable due to environment, resulting in a few erroneous identifications as high C18:1 - low C18:2 types, ie false positives if the latter was the desired type. The converse error seems less common. C18:3 percentage showed considerable overlap between the parents, and is only influenced in a minor manner by these genetics.

REFERENCES

1. Kirk, J.T.O. and Oram, R.N. 1981 Isolation of erucic acid free lines of Brassica juncea: Indian mustard now a potential oilseed crop in Australia. Journal of the Australian Institute of Science 47 51-52.

2. Love, H.K., Rakow, G., Raney, J.P. and Downey, R.K. 1990 Development of low glucosinolate mustard. Canadian Journal of Plant Science 70 419-424.

3. Seeds Regulations, Amendments to Consolidated Regulations of Canada, c 1400 (1987). Seeds other than seed potatoes, interpretation of ‘canola’. Canada Gazette Part II 121, 421.

¹ For F₁ plants, 1 at 10 seeds, 2 at 19 seeds and 1 at 34 seeds, otherwise 20 seeds per plant. For S₁ plants, 1 at 4 seeds, 12 at 10 seeds, and 1 at 21 seeds, otherwise 5 seeds per plant.