[Development of EST (Expressed Sequence Tags) Marker in Chinese Cabbage and its Transferability to Repeseed.].
28 pairs of primers were designed according to the expressed sequence tags in Chinese cabbage. After testing on the annealing temperature and the concentration of primer, dNTP and MgCl2, a suitable PCR system was established. Under the condition of reaction system developed, primers designed specific to ESTs were screened against genomic DNA of inbreed line A from which the cDNA library was constructed. Among them, 18 pairs of primers showed the amplification. Then all the primers available in line A were subjected to PCR for DNAs from 17 cabbage varieties. Polymorphism was detected by electrophoresis with agarose gel, and 10 of 18 primer sets could reveal polymorphisms among cabbage varieties, which accounted for 55.6% of primers selected. To examine the transferability of EST markers developed in cabbage, all primers were further used for PCR-mediated amplification of genomic DNA from different varieties of rapeseeds. Of 28 pairs of primers, 24 were able to produce amplified product(s) and 18 showed polymorphisms, accounting for 85.7% and 64.3% of total primers respectively. All of 18 primer sets that amplified in cabbage also showed amplified products in rapeseed and 13 of them were polymorphic. Even amongst the 10 primer sets that were unable to amplify in cabbage, 6 pairs produced amplification and 5 could reveal the polymorphism in rapeseeds. Results obtained in the present paper proved that developing polymorphic markers based on EST could be feasible and this kind of marker would be transferable to closed related species.
Institute of Muclear and Agricultural Sciences, College of Agriculture and Biotechnology, Zhejiang University, Hangzhou 310029, China; Email: xinya1015@yahoo.com.cn.
Ya
Y
Xin
Hai-Rui
HR
Cui
Ming-Long
ML
Zhang
Rong-Pyo
RP
Lin
Shui-Lian
SL
Cui
D001937:Brassica; D017931:DNA Primers / Q000235:genetics; D020224:Expressed Sequence Tags; D015723:Gene Library; D005819:Genetic Markers; D018895:Microsatellite Repeats; D011110:Polymorphism, Genetic
D017931:DNA Primers; D005819:Genetic Markers