Study of reproductive compatibility and morphological characterization of interspecific hybrids in Sesamum sp

In the present study, three wild species of sesame, Sesamum alatum, Sesamum malabaricum and Sesamum radiatum and one wild variety of Sesamum indicum, that is, S. indicum var. yanamalaiensis were crossed with eight cultivated varieties of S. indicum L. in both direct and reciprocal forms. All the wild species exhibited different degrees of cross compatibility with cultivated S. indicum. There was no crossed seed set in the direct and reciprocal crosses involving cultivars of S. indicum (2n = 26) with S. radiatum (2n = 64) and with S. alatum (2n = 26). The crosses involving S. malabaricum and S. indicum var. yanamalaiensis having the same chromosome number (2n = 26) as in the cultivated sesame genotypes were fairly successful in producing high percentage of crossed capsules with well filled seeds. The morphology of four wild species along with the cultivated species of sesame and the interspecific hybrids derived were compared. The wild species utilized in the present study differed significantly from the cultivated in branching pattern, leaf pubescence, flower size, color of corolla and anther, size, shape and color of extra floral nectary, capsule size, and shape, texture and size of the seed. All the successful interspecific hybrids showed predominance of wild characters than cultivated S. indicum.


INTRODUCTION
Sesame is known to be the most ancient oilseed crop dating back to 3050-3500 B.C. (Bedigian and Harlan, 1986) because of its ease of extraction, great stability, and drought resistance. It is also considered to be important because of its nutritional and antiaging features of high quality vegetable oil with oil content ranging from 50 to 60% (Chayjan, 2010). The sesame oil is highly resistant to oxidative deterioration due to the presence of antioxidants such as sesamin and sesamolin (Erbas et al., 2009) and also has high percentage of unsaturated *Corresponding to author. E-mail: meenacpbg_17@yahoo.co.in. Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0 International License fatty acids (Yermanos et al., 1972). Though sesame is having all these benefits, the productivity is limited due to low seed yield (Ashri 1989;Pham et al., 2010), frequent occurrence of diseases (El-Bramawy, 2006) and stress factors (Sarwar et al., 2007). Therefore, breeding efforts have mainly concentrated on increasing the seed yield of sesame. One of the important ways for increasing seed yield is utilization of diverse sources, especially the wild species for the exploitation of heterosis as well as to impart biotic and abiotic stress resistance. Hence an attempt was made to study the crossability between the four wild and cultivated species of sesame and to evaluate the hybrid vigour expression in the interspecific crosses.

MATERIALS AND METHODS
The experimental materials comprised of three wild species of sesame, Sesamum radiatum (2n = 64), Sesamum alatum (2n = 26), Sesamum malabaricum (2n = 26) and one wild form of Sesamum indicum, that is, S. indicum var. yanamalaiensis (2n = 26) as reported by Devarathinam and Sundaresan (1990) with eight cultivated varieties of S. indicum ( Figure 1). This includes CO 1, PYR 1, SVPR 1, VRI 1, VRI(Sv) 2, TMV 3, TMV 4 and TMV 7. The wild species were collected from the Species Garden maintained at Centre for Plant Breeding and Genetics, Tamil Nadu Agricultural University, Coimbatore, India. The varietal seeds were obtained from the Department of Oilseeds, TNAU, Coimbatore. The seedlings were raised in earthern pots during Summer, 2010. During flowering, the crossing was effected by utilizing the wild species as both male and female parents. This was done by emasculating the female flower buds (removing the corolla with four stamens) in the previous day evening. The just opened male flowers were collected from the respective parents on the following day morning and pollination was done between 6:30 to 8:30 AM. For this, the 1/3 rd of the corolla was removed to expose the stamen outside, which was then smeared on the stigma of the emasculated flower. A small paper tag was tied at the base of the pollinated flowers for easy identification of crossed capsules at the later stage. The entire crossing block was raised in the glass house to avoid insect pollination. The seeds were collected from the crossed capsules and the F 1 generation was raised with the parents, in two replications with each entry in two rows of 5 m length and spacing of 15 × 30 cm during Kharif, 2010. All the recommended package of practices was adopted.

Compatibility relationship
Based on the crossing data, it was evident that all the wild species exhibited different degrees of cross compatibility with cultivated S. indicum. The details about direct and reciprocal crosses attempted between the cultivars and the wild species are presented in Table 1. From the data on number of flowers pollinated, number of capsules set and number of hybrid seeds obtained, the crossability between the wild species and cultivated varieties were brought out. The overall cross compatibility relationship is given in Table 2.

Between S. indicum (2n = 26) and S. alatum (2n = 26)
Though S. indicum and S. alatum are having the same chromosome number, capsule and seed setting was not observed in both direct and reciprocal crosses of S. alatum with eight cultivars of S. indicum. Kedharnath (1954) could obtain only shriveled and non-viable seeds in S. indicum and S. alatum combination and presumed an early abortion of young embryo. Similar attempts were made by Amirtha Devarathnam (1965), Sundaram (1968) and Subramanian (1972) who also failed in producing viable hybrids between these two species. Even though these two species are having the same chromosome status (2n = 26), it is probable that a strong mechanism operates, due to which, hybrid seeds were not obtained.

Between S. indicum (2n = 26) and S. radiatum (2n = 64)
The direct crosses recorded the capsule setting with the maximum of 6.80% in S. radiatum × TMV 7 and minimum of 1.02% in S. radiatum × VRI 1. There was no seed set in any of the eight direct crosses between S. radiatum and S. indicum. The reciprocal cross between S. indicum and S. radiatum was not successful due to the premature dropping of crossed capsules. Hence, no capsule set and seed set was observed in the reciprocal crosses. Earlier studies involving these species also revealed the failure of these crosses (Dhawan, 1946;Ramanathan, 1950;Amirtha Devarathnam, 1965;Subramanian, 1972;Prabakaran, 1992;Vikas, 2006). The failure was attributed to very early collapse of the hybrid endosperm and the subsequent starvation of proembryo, as observed by Dhawan (1946) through embryological studies.

Between S. indicum (2n = 26) and S. malabaricum (2n = 26)
The crosses involving S. indicum cultivars and S. malabaricum both having the same somatic chromosome number (2n = 26) were fairly successful in producing good number of crossed capsules with well filled seeds. In the direct and reciprocal crosses effected between eight cultivars of S. indicum with S. malabaricum, successful capsule and seed setting was observed in all the 16 crosses ( Figure 2). Only one crossed seed got germinated in the cross between S. malabaricum as female with S. indicum genotype SVPR 1, but the seedling was died subsequently in the two leaves stage, probably due to the abiotic factors. In the earlier description by John et al. (1950), S. malabaricum was referred as the variety of S. indicum, as S. indicum var. malabaricum, which was highly compatible with other genotypes of S. indicum. However, Prabakaran (1992)   referred this as the separate species of sesame as S. malabaricum. He reported that S. malabaricum had possessed distinct morphological features like longer duration, green stem with purple tinge, leathery leaves, purple corolla, highly rough testa as seen against the cultivated sesame. Also, S. malabaricum had shown partial capsule set when crossed with cultivated S. indicum (Prabakaran, 1992). The percentage of capsule setting ranged from 4.6% (S. malabaricum × CO 1) to 7.9% (S. malabaricum × VRI 1) in direct crosses. In reciprocal crosses, it was between 0.9% (TMV 3 × S. malabaricum) and 12.6% (SVPR 1 × S. malabaricum).
In direct crosses, the mean number of seeds per capsule was the lowest in S. malabaricum × SVPR 1 (6.7) and highest in S. malabaricum × TMV 7 (25.7). Similarly, the cross SVPR 1 × S. malabaricum recorded the lowest number of seeds per capsule (13.1) and the highest was recorded in TMV 7 × S. malabaricum (55.1) in reciprocal crosses.

Between S. indicum (2n = 26) and S. indicum var. yanamalaiensis(2n = 26)
The cross-compatibility between S. indicum and S. indicum var. yanamalaiensis both having the same chromosome number of 2n = 26 was confirmed both in direct and reciprocal form (Figure 2). But the capsule set and seed set was not observed in S. indicum var. yanamalaiensis with TMV 3 and TMV 4. The range of capsule setting was from 0 to 52.2% in S. indicum var. yanamalaiensis × VRI(Sv) 2. The seed setting was ranged from 0 to 22.3% (S. indicum var. yanamalaiensis × Paiyur 1) in direct crosses. In the cross S. indicum var. yanamalaiensis × VRI 1, crossed seed was obtained but the seeds were small and shriveled and hence not germinated.
In the reciprocal crosses, there was capsule set, but no seed set in TMV 4 × S. indicum var. yanamalaiensis. The range of capsule setting was from 1.0% (TMV 3 × S. indicum var. yanamalaiensis) to 14.1% in SVPR 1 × S. indicum var. yanamalaiensis. The seed setting was ranged from 0 (TMV 4 × S. indicum var. yanamalaiensis) to 47.5% (TMV 7 × S. indicum var. yanamalaiensis). Since the flowering of both parents had not coincided and hence, the pollination was attempted in the later stage of flowering. Due to this, the seed set was not observed in few of the direct and reciprocal crosses between S. indicum var. yanamalaiensis and cultivated varieties.

Parents
The morphology of four wild species and the cultivated species of sesame was compared and given in Table 3. The wild species utilized in the present study differed significantly from cultivated one in the branching pattern, leaf pubescence, flower size, color of corolla and anther, size, shape and color of extra floral nectary and capsules, texture and size of the seed. S. alatum was profusely branching with completely lobed basal leaves. The corolla color was maroon and glabrous with dark purple corolla lip. The anther was dark purple with purple colored extra floral nectary. The capsules were long and tapering with small and winged seeds. The branches of S. malabaricum were profuse with pubescent leaves. The Kumari and Ganesamurthy 915 corolla was pink and densely hairy with dark pink colored corolla lip. The calyx also had dense hairs with flower having purple anther. The glands were yellow colored and prominent. The capsules were medium sized and hairy. The seeds were also medium sized with rough testa. S. indicum var. yanamalaiensis resembled cultivated S. indicum in most of the traits. It differed from cultivars in branching pattern, corolla and corolla lip color and in the size of yellow glands. The capsules were medium sized sparsely hairy with small black colored seeds with smooth testa as in the cultivated varieties. The wild species S. radiatum differed widely from S. indicum. The stem of S. radiatum was pubescent with more number of branches. The leaves were dark green, pubescent with serrated margins. The corolla was hairy, purple colored with dark purple corolla lip. The calyx was also pubescent with flowers having big, cream colored anther. The glands were dark colored with densely hairy capsules. The seeds were small with rough testa. These above mentioned specific traits were not observed in the cultivated S. indicum genotypes.

Inter-specific hybrids
The observed morphological characters of the direct and reciprocal crosses of wild with cultivated species are given in Table 4. The hybrids developed from the direct and reciprocal crosses involving S. malabaricum and S. indicum were similar in the expression of qualitative traits. But, the hybrids with S. malabaricum as the female parent had taken comparatively more days to germinate, when compared to their reciprocals. This difference was due to the maternal seed traits of the wild parent. The duration taken for germination of hybrids is much more than their cultivar parent.
The hybrids exhibited most of the phenotypic characters of wild parent, indicating the dominant nature of S. malabaricum. The direct crosses resembled the wild parent, S. malabaricum in branching pattern, leaf pubescence, corolla and corolla lip color, flower having calyx with dense hairs, and light purple colored anther.
The nature and color of extra floral nectary resembled the wild parent. The capsules were very small with few seeds, which was medium sized black with rough testa. The crossed seeds had expressed dormancy as in the wild parent and many of the crossed seeds had not germinated for more than two months. The reciprocal crosses had also expressed similar traits as in direct crosses between S. malabaricum and S. indicum.
The F 1 hybrids involving the eight cultivars of S. indicum with S. indicum var. yanamalaiensis, were evaluated for their morphology and it was found that the 12 hybrids resembled the wild parent in branching pattern, corolla and corolla lip color. From this study, it was found that all the successful interspecific hybrids    showed predominance of wild characters than cultivated S. indicum. The wild species viz., S. malabaricum and S. indicum var. yanamalaiensis could be effectively utilized for the transfer of essential traits from wild to cultivated through conventional breeding program.