| Castor (Ricinus communis L.) is a traditional oil plant which has special purpose and high economic value. It is one of top-ten important oil crops of world, The traditional breeding methods can only select from the phenotype not from molecular level and it costs too much time and energy. To utilize the advantage of hybrid breed is an effective way to improve the castor's output and quality. Castor Hybridizing mainly from the use of pistillate castor, the discovery and use of pistillate castor opens up a new way to hybridize breeding. Very few research about pistillate characters in castor breeding. In this paper, if a molecular marker with close linked to castor pistillate character had been obtained, we can select pure female character fast and accurately, shorten breeding time and accelerate the acquisition of female strains.The main results were as follows:1. The discussion of genomic DNA extraction from castorGenomic DNA was extracted by modified CTAB method and SDS method. Agarose gel electrophoresis showed that the modified CTAB method could obtain higher quality genomic DNA with better purity and integrity compared with SDS method. Castor's leaves contain abundant polysaccharide and polyphenols, so a proper amount of P-mercaptoethanol and PVP were added in order to prevent oxidation of phenolic compounds.2. One set of stable ISSR reaction system was establishedThe influences to ISSR-PCR were optimized were tested using single factor experiment. A reaction system suitable for pistillate castor were established, that is 20μL amplification reactions system containing 10×PCR buffer 2.5μL,10 ng template DNA,2.0 mmol/L MgCl2,1.0μmol/L primer, 0.4 mmol/L dNTPs,1.0 U Taq DNA polymerase。3. Screening of specific ISSR primers According to BSA method, two gene pools were made up:pistillate pool and amphoteric pool. Through amplify repeat, it showed that primer UBC844 could amplified a special band in female plant and pistillate pool, and its length was about 750bp. By the further test with individuals of F2 segregation population, primer UBC844 amplified a stable band UBC844750, which was found in most of pistillate plants, but was absent in amphoteric plants. It showed that UBC844750 may be close linked with castor pistillate character. |
