Sorghum [Sorghum bicolor (L.) Moench] is ranked the fifth most important cereal crop in the world. The United States, India, Nigeria, Mexico, Sudan, and China currently produce the most grain sorghum. More than half the world's sorghum is grown in the semi-arid tropics, where it is a staple food for millions of people in India and Africa; however, livestock feeding accounts for most of the U.S. sorghum usage.

 

Many studies have been devoted to assessing patterns of sorghum genetic variation based on morphology (Appa-Rao et al., 1996; Djè et al., 1998) or pedigree (Jordan et al., 1998). More recently, DNA-based techniques have been used successfully in DNA fingerprinting of plant genomes (Hongtrakul et al., 1997; Cervera et al., 1998) and in genetic diversity studies (Paul et al., 1997; Sonnate et al., 1997; Barrett and Kidwell, 1998; Chowdari et al., 1998b; Zhu et al., 1998; De-Bustos et al., 1999). 

 

Among them, random amplified polymorphic DNA (RAPD) analysis is quick (Colombo et al., 1998; Fahima et al., 1999) and well adapted for nonradioactive DNA fingerprinting of genotypes (Cao et al., 1999). However, problems with the reproducibility in amplification of RAPD markers and with data scoring have been reported (Jones et al., 1998). Although major bands from RAPD reactions are highly reproducible, minor bands can be difficulty to repeat due to the random priming nature of this PCR reaction and potential confounding effects associated with co-migration with other markers (Tessier et al., 1999).

 

SSR markers are attractive for DNA fingerprinting studies for several reasons. They are codominant and highly informative. They generally display high levels of polymorphism (Beckmann and Soller, 1990; Brown et al., 1996; Senior et al., 1998) and are amenable to automated genotyping strategies. They also can be amplified by PCR and efficiently detect DNA polymorphism (Pejic et al., 1998). Finally, radioisotopes are not required in the detection of SSR markers, because sequence polymorphism usually can be detected by separation in agarose gels (Burr, 1994).

 

Although SSRs are well established for human and mammalian genetics, these markers have only recently become available in plant species. They have been identified in many plant genomes including those of maize (Senior and Heun, 1993; Shatuck-Eidens et al., 1990; Taramino and Tingey, 1996); soybean (Akkaya et al., 1992; Morgante and Olivieri, 1993); Brassica spp. (Poulsen et al., 1993); rice (Wu and Tanksley 1993); barley (Saghai-Maroof et al., 1994); pearl millet (Chowdari et al., 1998a); Arabidopsis (Depeige et al., 1995); tomato (Broun and Tanksley, 1996); conifers (Tsumura et al., 1997); and sorghum (Brown et al., 1996; Taramino et al., 1997; Dean et al., 1999). The results of studies using SSR markers in these species suggest that they may provide an outstanding new tool for genetic analysis of plant species.

 

Harlan and DeWet (1972) classified cultivated sorghum based on agronomic and morphological characteristics. The utilities of isozymes (Morden et al., 1989; Aldrich et al., 1992), RFLP (Aldrich and Doebley, 1992), and RAPD (de Oliveira et al., 1996; Menkir et al., 1997; Ayana et al., 2000) markers have been used to study genetic diversity in sorghum germplasm. Several efforts have been made to utilize SSR markers in plants to study genetic diversity, characterize germplasm, and evaluate population dynamics (Zhang et al., 1997; Liu and Wu 1998; Senior et al., 1998; Struss and Plieske, 1998). A comparison of RAPD and SSR marker techniques in sorghum is timely, even though the utility of different molecular markers for corn (Smith and Helentjaris, 1996), soybeans (Powell et al., 1996) and barley (Russell et al., 1997) germplasm already has been reported. The objectives of the present study were to: (1) compare the application and utility of RAPD and SSR marker techniques for analysis of genetic diversity among sorghum genotypes, (2) compare genetic similarity quantified by molecular markers with regional and race information.

 

 

Plant Materials

 

Twenty-two sorghum accessions including landraces, improved lines, and wild accessions were evaluated in this study (Table 1). Most of these accessions represent landrace varieties and are described in detail in the USDA-ARS Germplasm Resources Information Network available at http://www.ars-grin.gov/npgs. Less information is available on wild accessions and improved varieties that were obtained from plant breeders from different parts of the world.

   

 

Table 1. Country of origin, race classification, and other distinguishing characteristics for sorghum accessions used in this genetic diversity studies.

 

No	Germplasm Accession	GRIN Designation	Origin	Region	Race	Distinguishing Characteristics
1	P954035 (SC 33)	PI 534132	Ethiopia	1	D	Drought tolerance
2	B35 (SC 35)	PI 534133	Ethiopia	1	D	Drought tolerance
3	SC 1158	PI 597957	Ethiopia	1	D	Disease resistance
4	SC 326	IS 3758C	Ethiopia	1	C	Disease resistance
5	SC 414	PI 533831	Sudan	2	C	Disease resistance
6	SC599	PI 534163	Sudan	2	C	Drought tolerance
7	ShanQuiRed	-	China	7	B	Cold tolerance
8	SanChiSan	GRIF 620	China	7	B	Cold tolerance
9	PI 550590	PI 550590	Russia	8	B	Cold tolerance
10	12-26	-	Egypt	5	W	Wild sorghum
11	47-121	-	Kenya	1	W	Wild sorghum
12	PI 465483	PI 465483	Yemen	1	D	Large seed size
13	PI 559761	PI 559761	Yemen	1	D	Large seed size
14	Dorado	-	El-Salvador	6	M	Elite tropical variety
15	Malisor	-	Mali	3	M	Elite tropical variety
16	Sureno	-	Honduras	6	M	Elite tropical variety
17	Macia	-	Mozambique	4	M	Elite tropical variety
18	M91051	-	USA	6	M	Elite tropical variety
19	TX430	PL-140	USA	6	M	Elite U.S. pollinator
20	TX2737	GP-82	USA	6	M	Elite U.S. pollinator
21	TX2741	GP-86	USA	6	M	Elite U.S. pollinator
22	PL1	-	USA	6	M	Large seed size

 

Region: 1 = East Africa, 2 = Central Africa, 3 = West Africa, 4 = South Africa, 5 = Northern Africa, 6 = North America, 7 = Asia, 8 = Europe. Race: D = Durra, C = Caudatum, B = Bicolor, W = Wild, M = Breeding.

 

 

DNA extraction and SSR and RAPD markers

 

Genomic DNA was extracted from etiolated hypocotyls 5- to 7-day-old plants of each genotype according to the method of Djè et al. (2000). Initially the five individual plants of each accession were assayed for RAPDs or SSRs using 5 primers. No polymorphisms were detected between individuals within a genotype.

 

Seventeen SSR markers described by Brown et al. (1996) and 11 described by Taramino et al. (1997) were used for genotyping assays (Table 2). Eighty different RAPD primers obtained from OPERON Technologies (Kits A-D) were used to generate markers as described by Tao et al. (1993). Thirty-two primers that generated clear and reproducible fragments were used to fingerprint the 22 sorghum genotypes. The SSR and RAPD reaction products were evaluated for polymorphisms on 3% Metaphor agarose gels (FMC Products, Rockland, ME, USA) and 1.6% agarose gels, respectively. Gels were stained with 1 mg mL^-1 ethidium bromide for 30 to 60 min.

 

   

                        Table 2. SSR marker used to diverse the 22 sorghum accessions.