Research Article: Development of novel SSR markers for evaluation of genetic diversity and population structure in Tribulus terrestris L. (Zygophyllaceae)

Date Published: July 19, 2016

Publisher: Springer Berlin Heidelberg

Author(s): Kuljit Kaur, Vikas Sharma, Vijay Singh, Mohammad Saleem Wani, Raghbir Chand Gupta.


Tribulus terrestris L., commonly called puncture vine and gokhru, is an important member of Zygophyllaceae. The species is highly important in context to therapeutic uses and provides important active principles responsible for treatment of various diseases and also used as tonic. It is widely distributed in tropical regions of India and the world. However, status of its genetic diversity remained concealed due to lack of research work in this species. In present study, genetic diversity and structure of different populations of T. terrestris from north India was examined at molecular level using newly developed Simple Sequence Repeat (SSR) markers. In total, 20 primers produced 48 alleles in a size range of 100–500 bp with maximum (4) fragments amplified by TTMS-1, TTMS-25 and TTMS-33. Mean Polymorphism Information Content (PIC) and Marker Index (MI) were 0.368 and 1.01, respectively. Dendrogram showed three groups, one of which was purely containing accessions from Rajasthan while other two groups corresponded to Punjab and Haryana regions with intermixing of few other accessions. Analysis of molecular variance partitioned 76 % genetic variance within populations and 24 % among populations. Bayesian model based STRUCTURE analysis detected two genetic stocks for analyzed germplasm and also detected some admixed individuals. Different geographical populations of this species showed high level of genetic diversity. Results of present study can be useful in identifying diverse accessions and management of this plant resource. Moreover, the novel SSR markers developed can be utilized for various genetic analyses in this species in future.

Partial Text

Tribulus terrestris commonly known as puncture vine and Gokhru, belonging to family Zygophyllaceae, is an annual herbaceous plant. The plant is native to the South and East Europe and West Asia. It is widely distributed in the warm regions of Asia, Africa, Europe, America and Australia (Topia et al. 1994; Abeywickrama and Bean 1991; Kostova et al. 2002). The species is commonly distributed throughout tropical and warmer regions of India. It occurs naturally in many Indian states with warm climate and reported from eastern, western, northern, southern and central parts of country (Mishra and Bisht 2012; Das and Ghosh 2014; Fatima et al. 2014; Pandey 2014, 2015). It is a prostrate to procumbent annual, hairy herb. Leaves are pinnately compound, leaflets are 4–8 paired, subsessile, ovate or elliptic. Flowers are yellowish colored, mericarps not winged but distinctly spinous (Fig. 1). The fruits of the species are very distinguished in nature like a stellate and are known as ‘Chih-hsing’ in China and ‘Goat head’ in USA. It is used as traditional ayurvedic medicine in various health disorders. It contains saponins, steroids, estradiol, flavonoids, alkaloids, unsaturated fatty acids, vitamins, tannins, resins, nitrate potassium, aspartic acid and glutamic acid (Gauthaman and Adaikan 2005). The plant is a rich source of saponins, of which protodioscin has received a large attention in regards to sexual dysfunction issues (Adimoelja 2000). Now-a-days, full plant or its fruits are used in large number of purposes as skin-care, human hormone regulation, antibacterial, anti-inflammation, antivirus and immunostimulant. The whole plant is useful in strangury, dyspepsia, helminthiasis, cough, asthma, cardiopathy, skin diseases, hypertension and rheumatic arthritis (Sivarajan and Balachandran 1994; Warrier et al. 1996; Petkov 2011). Thus, it is well known that all parts of the plant have great medicinal potential. T. terrestris exhibits both, self- and cross-pollination mechanisms (Ganie 2011). This herb propagates through the seeds only. The species is dibasic depending on base number x = 6 and 10. Cytologically, the species is quite variable with intra-specific polyploids from diploid to octaploid (2n = 12, 24, 36, 48; Morrison and Scott 1996). From India, the chromosome count of 2n = 24, 32, 36, 48, 72, 96 has been reported by various workers. From Rajasthan, three cytotypes of the species, i.e., tetraploid, hexaploid and octaploid have been reported (Gupta et al. 2016). Morrison and Scott (1996) suggested that these various polpyploids are originated from allopolyploid complex. Propagation through seeds presents opportunity for analyzing polymorphisms of these populations which are not so far apart, as reproductive mode results in allelic recombinations. In northern parts of India, this species is widely found in Punjab, Haryana and Rajasthan. As it requires very less water for its growth, it is commonly found in bare and uncultivated lands. Although, T. terrestris is cultivated in various regions of Rajasthan for its medicinal utilities but these practices further needs elite germplasm for sustainable utilization. On the other, due to preference to major cereal crops and other cash crops in Punjab and Haryana, populations of this species often uprooted in large scale which is posing a threat to genetic diversity of this species. Therefore, it becomes imperative to characterize its existing germplasm so that diverse germplasm can be identified and maintained for future.Fig. 1a A photo of plant at natural habitat of Punjab showing plant bearing leaves and flowers. b Enlarged photo of flower. c Seeds of T. terrestris

In conclusion, this was the first attempt of SSR development in T. terrestris and these markers appeared highly polymorphic and informative in characterized germplasm. Moreover, genetic diversity and population structure related studies are severely lacking in this species. Therefore, the novel information and SSR markers provided here can be useful in accelerating these types of studies in different germplasms of T. terrestris. We found that there is considerable genetic diversity prevailing in north Indian germplasm of T. terrestris which needs preservation can be exploited in a sustainable manner. The results of present study can be helpful in planning the utilization patterns and management of T. terrestris germplasm existing in north India. The novel SSR markers developed in this study can be useful in future genetic characterization related studies of the germplasm of this species.




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