1.College of Life Science, Fujian Agriculture and Forestry University, Fuzhou,China 350002,2. Key Laboratory of Pesticide and Chemical Biology, Ministry of Education, China
Random amplified polymorphic DNA (RAPD) markers were used to estimating genetic diversity among 57 allelopathic rice accessions (Oryza sativa L.). There were 59 polymorphic bands in the 85 RAPD loci within the tested accessions, and the percentage of polymorphic bands (PPB) was 69.4%. The result from the clustering analysis by Unweighted Pair Group Mathod Arithmetic Average (UMGMA) indicated that those accessions from the same geographical location were clustered into one group. It was also found that some rice accessions with higher allelopathic potential were clustered together, such as IAC25、IAC47and IAC120 implying that the genes conferring allelopathy in those rice accessions might be isolocus. However, some rice accessions with different allelopathic potential clustered into the same group performed lower level of generic polymorphism which was attributed to oriented selection for other traits in breeding program. This offers us a genetic pool for the selection of rice cultivars with high allelopathic ability.
Key Words
Allelopahic rice accessions; Oryza sativa L., Genetic diversity; RAPD
Weeds are one of the most important causes of yields losses in rice. In the upland rice ecosystem, yield loss estimates from weed infestation have ranged from 30 to 100% (Luoise 2001). With the increasing of labor-cost increasing and the availability and efficacy of herbicide, herbicide use has increased exponentially in Asian rice production (Naylor 1996).Despite the overwhelming economic benefit, the heavy reliance on chemical herbicides is considered a potential threat to public health and to the environment (Jiwan and Gates,1994).Furthermore, the appearance of herbicides-resistance weeds is an increasing problem (Boutsalis and Powles,1995).These factors have prompted research into alternative means of weed control.( Motiul Quader et al,2001).
Allelopathy is defined as “direct or indirect (harmful or beneficial) effects of a plant, on another plant through the release of compounds that escapes into the environment”(Rice,1984).Three seasons of field experience finally led to the conclusion that allelopathy could explain 34% of the variation among cultivars in the ability to suppress weeds (Olofsdotter et al.1999).
Allelopathic potential has been identified in most major cereal crops (Lovett and Hoult,1995).Based on the selection for rice accessions , the assessment of genetic diversity became necessary when a significant level of intraaccession variability in allelopathic expression became evident ( Motiul Quader et al,2001).
Fifty-seven rice accessions of Oryza sativa L. introduced from American, Korea, Brazil, China and the International Rice Research Institute (IRRI), were screened for allelopathic potential. Barnyardgrass (Echinochloa crus-galli L.)was used as a receiver plant.
The procedure was adapted from the relay seedling technique (Navarez and Olofedotter 1996). 0.5% water agar was used as growth medium after compared with Perlite (the data unpublished). The root length of barnyardgrass was recorded.
All seeds were grown in Petri dishes containing 0.5% water agar under sterile conditions. Genomic DNA were extracted from duplicate samples from a single seedling by the method of Weining et al.(1994). The polymerase chain reaction (PCR) was performed in a 15μl reaction mixture containing 10ng template DNA,1.5μl 10×buffer,200μmol/L dNTPs,1.5μl Taq DNA polymerase and 1ng 10-mer primer. Amplification program is :30s at 94° ;30s at 40° ;90s at 72° ;40 cycles, 10min at 72° then stored at 4°。
The root length of barnyardgrass was transferred into Inhibition Ratio (IR), i.e. IR=(1-TR/CK)×100%, while TR represented the treatment,CK was the control.
Genetic data analyses were performed by using the computer package TFPMG 1.3(Miller et al.1997).The dendrograms was constructed by the unweighted pair group method (UPGMA)(Sneath and Sokal,1973).
Of 57 accessions, 5 rice cultivars, such as Iguape Cateto, PI312777, Azucena, Taichung Native 1 and IAC25, demonstrated over 50% inhibition of barnyardgrass root growth. IRs of 12 cultivars ranged from 40% to 50%, that of 21 cultivars from 30% to 40%,13 cultivars from 20% to 30%, while IRs of 6 cultivars were less than 20%(Table 1). The distribution of allelopathic activity in rice accessions tested was normal(Fig 1).
Table 1: IRs of rice alleloapthy on the root of barnyardgrass*
No |
Rice accession |
Origin |
IR (%) |
No |
Rice accession |
Origin |
IR (%) |
1 |
Iguape Cateto |
Brazil |
58.4±1.6 |
30 |
Chaoerzhan |
China |
33.3±6.3 |
2 |
PI312777 |
America |
56.8±2..3 |
31 |
Sanyizhaozhan |
China |
32.7±6.7 |
3 |
Azucena |
The Philippines |
53.9±1.4 |
32 |
Wab56-125 |
Ivory Coast |
32.5±4.1 |
4 |
Taichung Native 1 |
Taiwan |
50.2±8.9 |
33 |
Polha Murcha |
Brazil |
32.0±6.6 |
5 |
IAC25 |
Brazil |
50.0±4.7 |
34 |
Qisanzhan |
China |
31.8±7.5 |
6 |
AU257 |
Bangladesh |
48.4±0.6 |
35 |
Dourado Pecoce |
Brazil |
31.8±3.5 |
7 |
Red Rice5 |
China |
48.2±6.8 |
36 |
Bala |
India |
31.6±6.7 |
8 |
Batatais |
Brazil |
47.5±5.2 |
37 |
Arroz de campos |
Cuba |
30.7±7.9 |
9 |
IAC120 |
Brazil |
46.7±7.4 |
38 |
Shuangzhan 2 |
China |
30.4±7.5 |
10 |
Co39 |
India |
45.2±2.6 |
39 |
Fengaizhan |
China |
29.1±5.8 |
11 |
IAC47 |
Brazil |
45.1±7.5 |
40 |
IR721413 |
The Philippines |
28.9±4.7 |
12 |
IR72417-3R-8-2 |
The Philippines |
44.5±13.2 |
41 |
Qidaizhan |
China |
28.9±8.8 |
13 |
Yehuazhan |
China |
43.6±5.9 |
42 |
IR73384 |
The Philippines |
27.7±9.4 |
14 |
IR70617 |
The Philippines |
43.2±6.7 |
43 |
IR64 |
The Philippines |
27.7±5.3 |
15 |
Jingyouzhan |
China |
41.1±8.6 |
44 |
Xinsimiao |
China |
27.3±1.3 |
16 |
IAC164 |
Brazil |
41.1±6.9 |
45 |
Daishuzhan |
China |
27.0±4.3 |
17 |
Mafeng 1 |
China |
40.9±10.1 |
46 |
Qingxiangzhan |
China |
26.8±6.1 |
18 |
189Taizhong 189 |
China |
39.3±8.6 |
47 |
IR62266-42-6-2 |
The Philippines |
25.9±4.0 |
19 |
Dinorado |
The Philippines |
39.3±2.2 |
48 |
IR65907-116-1-B |
The Philippines |
24.1±9.1 |
20 |
Vandana |
Colombia |
38.2±5.6 |
49 |
Moroberekan |
Guinea |
23.2±6.6 |
21 |
IAC165 |
Brazil |
37.5±1.1 |
50 |
IR60080-46A |
The Philippines |
21.4±4.1 |
22 |
IR56 |
The Philippines |
36.6±4.7 |
51 |
IR72412 |
The Philippines |
20.2±7.8 |
23 |
Shuangmeizhan |
China |
36.4±6.4 |
52 |
Zhengyou 1 |
China |
19.5±1.9 |
24 |
IR70651 |
The Philippines |
35.7±2.4 |
53 |
IR55423-01 |
The Philippines |
18.4±2.5 |
25 |
IR36 |
The Philippines |
34.5±4.3 |
54 |
Pratao Precoce |
Brazil |
18.2±2.6 |
26 |
Dee Geo Woo Gen |
Taiwan |
34.3±3.8 |
55 |
Aisanruzhan |
China |
17.7±4.6 |
27 |
Muxiang 25 |
China |
34.1±3.1 |
56 |
Dular |
America |
14.1±2.1 |
28 |
IR73382 |
The Philippines |
33.9±2.8 |
57 |
Lemont |
America |
10.9±3.2 |
29 |
IR71331 |
The Philippines |
33.3±9.4 |
CK |
0.0 |
*CK, Control= Echinochloa crusgalli L. grown alone
Fig.1(Left) The distribution proportion of rice accessions in different interval of IR value
Fig.2 (Right) The cluster analysis on allelopathic rice as determined by UPGMA from RAPD
12 primers were selected from a previous study on the basis of polymorphisms revealed in 5 rice accessions. The primers were used to detect the polymorphism of 57 rice accessions (Figure 3). The total of 85 bands were scorable, of which 59 were polymorphic, and the percentage of polymorphic bands(PPB) was 69.4%.
Fig.3 DNA fragment amplified by primer 1409
From GD=0.1875, 57 rice accessions were grouped into eight main group(Fig.2). The first group was composed of 31 cultivars, of which 14 were from Mainland China, 13 from the Philippines, 2 from Taiwan, 1 from American and 1 from India. In this group, 8 cultivars showed over 40% of inhibitory effect on the root of baryardgrass. The sixth group included 6 accessions introduced from Brazil. Three cultivars, IAC25(5), IAC47(11) and IAC120(9), which all showed high allelopathic potential on barnyardgrass, were clustered together. Another three cultivars, Iguape Cateto(1), Dourado Pecoce(35) and Pratao Precoce(54) were grouped into one subgroup, performing that the inhibition ratio (IR) were 58.4%, 31.8% and 18.2% respectively.
In this study, the genetic polymorphism of allelopathic rice detected by RAPD approach indicated that those accessions from the same geographical location were clustered into one group. It was also found that some rice accessions with higher allelopathic potential were clustered together, implying that the genes conferring allelopathy in those rice accessions might be isolocus. However, some rice accessions with different allelopathic potential clustered into the same group, such as Iguape Cateto(1), Dourado Pecoce(35) and Pratao Precoce(54) , performed lower level of generic polymorphism which was attributed to oriented selection for other traits in breeding program. Bustos et al (1998) have successfully used RAPD markers to investigate the genetic diversity within and among wild populations of species of the genus Hordeum (Poaceae). With careful screening and replication, M.Quader et al (2001) observed high level of intraaccession genetic diversity (4-24%) in accessions of Triticum Speltoides by using RAPD.
This work was supported by grants (30200170, 2003F012) from National Natural Science Foundation of China, and The key Scientific Technological Program of Fujian Province, China.
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