| The soybean is one of the main crops in China, being the important food, animal feed and the industry raw materials.In soybean production, the plant diseases and insect pests is one of the important factors .Therefore, how to control plant diseases and insect pests effectively,having been important problem that the soybean production must face. Phytophthora rot causes by Phytophthora sojae.This disease was observed in Indiana in 1948. Since that time, Phytophthora rot has been reported in Argentina,Australia,Brazil,Canada,the People's Republic of China,Hungary,Italy,Japan,the former Soviet Union,and throughout the soybean-growing regions of the United States.In this study, it aims at mining SSR markers from ESTs of P.sojae, designing primer based on these EST-SSRs.The genetic characteristic and diversity in population of P.sojae and other fungus was evaluated are analyzed by EST-SSRs DNA fingerprint. Meanwhile, the research of the rapid detection of P.sojae which are carried by soybean is done by this method. The following are the main research methods and results of the study:1, Mining SSR from 28197 ESTs of P.sojae which total length is 9.96Mb by using SSRIT. 1574 EST-SSRs are found. In these ESTs, 1346 ESTs have one SSR(4.77%),108 ESTs have two and more SSRs(0.38%).Among these SSRs, most of these are Trinucleotide, Trinucleotide have 855 and is54.32% of total numbers.Hexanucleotide have296(18.81%) Mononucleotide have 125(7.94%) ,Pentanucleotide have121(7.69%),Tetranucleotide have 100(6.35%),Dinucleotide have77( 4.89%).2> Selected the P.sojae EST- SSRs to design and synthesized 243 pairs ofPrimer. These primers are detected by PCR amplify which using 10different P.sojae DNA samples.193 pairs primer have functionC79.4%). 120amplifier products are single strip(62.2%) and 73 amplifier products aremuliti strips (37.8%).3> Study on the transferability of P.sojae EST- SSRs on soybean and otherfungus. Selected 44 pairs primer which amplifier products are mulitistrips.26 pairs primer can amplify on soybean DNA samples.4n Selecting 29 SSR markers to analyses the relationships of P.sojae, P.Infestans, Pythium, P. nicotianae, P. citrophthora, P. boehmeriae, P.cactorum, P. cinnamomi, P. insolita, P. palmivora, P. crypotopea,tic. fourdifferent P. nicotianae samples are togather,two P. cactorum samples aretogether. P.sojae and P. Infestans can't assemble with other fungus.5^ Used 51 different P.sojae DNA samples,48 other fungus DNA samplesand two soybean DNA samples which successfully PCR amplified usingprimers to rapid detect P.sojae. Selected two pairs of primer which P.sojaeDNA amplifier products can separate with other fungus DNA amplifier.6^ Among 37 pairs functional primer.selected 23 pairs of primer whichamplifier products are single strip and 13 pairs of primer which amplifierproducts are muliti strips by using 51 different P.sojae DNA samples.7^ Genetic variation 51 P.sojae which isolates from different places andsources,was analyzed by SSR marker assay.Using 32 pairs of primer,52 fingerprint patterns were generated.The results showed that substantial genetic diversity existed among isolates.Most of American islates are together with Hei longj iang'isloates.as follows: PSUSA17 gather with PSJ99-8 and PSCJ99-1, PSUSA29 gather with PSJ21, PSUSA31 and PSUSA28 gather with PSJ99-13, PSUSA30 gather withPSJNK16,PSAMR gather with PSJMSl.But, Anhui'islate- PSCMC3 and Brazilian islate-PSBX-1 can't gather with other isolates.The relationship between Chinese islates and American islates is very near,and Hei long jiang'islates may come from America. |
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