Analysis on DNA methylation of mosaic and moxa type leaves of Aconitum carmichaeli by MSAP / 中国中药杂志

An MSAP analysis method was established for detecting DNA methylation of Aconitum carmichaeli leaves, and the DNA methylation of different leaf shapes and different leaf position was analyzed by MSAP. The study made experiments on the leaves of different position of mosaic and moxa leaf type A. carmichaeli, researched the effects of restriction digestion of genomic DNA by using two restriction enzymes, screened the suitable selective amplification primers, and analyzed the methylation differences of leaves by calculating the 6% acrylamide gel electrophoresis bands and lane. The best reaction system of MSAP was obtained, under the conditions of 37 ℃, the 16 h incubated time was more suitable for 150 ng DNA, and 25 pairs of selective amplification primers were selected from 256 pairs. Totally, 273 electrophoresis bands were obtained by 25 pairs of selective primers, including 228 non methylation or single chain methylation bands,27 double chain methylation bands,and 18 single stranded methylation bands, the total methylation rate was 16.48%. The methylation rate was slightly different in mosaic and moxa leaf type A. carmichaeli leaf, which were 15.36%, 14.34%, respectively, and article 8, article 6 nucleotide fragments of genome methylation modification differences were obtained, accounted for 3%, 2.26% of the total number of bands. Based on this study it can provide new ideas for molecular identification, breeding and cultivation, and genetic evolution of A. carmichaeli.

Effect of nickel chloride on Arabidopsis genomic DNA and methylation of 18S rDNA

Background In recent years, nickel (Ni) has been widely applied in industrial and agricultural production and has become a kind of environmental pollution. In this study, the effect of nickel chloride (NiCl2) with different concentrations on Arabidopsis genomic stability and DNA methylation has been demonstrated. The nucleolus variation and 18S rDNA methylation after NiCl2 treatment have been analyzed. Results The results are as follows: (1) The NiCl2 could result in heritable genomic methylation variations. The genomic DNA methylation variations have been detected by methylation-sensitive amplified polymorphism (MSAP) molecular markers, and the result showed that after NiCl2 treatment, there was methylation variation in T0 generation seedlings, and partial site changes maintained in T1 generation, which suggested that the effects of NiCl2 on DNA methylation could be heritable in offspring. (2) NiCl2 brought deformity and damage to nucleolar structure in Arabidopsis root tip cells, and the damage was positively correlated with the NiCl2 concentration. 3. In the nucleolus, there was an increased cytosine methylation in 18S rDNA. The plant nucleolus variation and 18S rDNA methylation may be used as an examination indicator for Ni pollution in soil or plant. Conclusions NiCl2 application caused variation of DNA methylation of the Arabidopsis genomic and offspring's. NiCl2 also resulted in nucleolar injury and deformity of root tip cells. The methylation rate of 18S rDNA also changed by adding NiCl2.

DNA Methylation of polyploid complex of Pinellia ternata by MSAP analysis / 中草药

Objective To analyze the DNA methylation status of polyploid complex of Pinellia ternata. Methods Methylation sensitive amplified polymorphism(MSAP) technique was carried out to analyze the methylation status of polyploid complex of P.ternata. Thirty-four selective primer amplifications were used to check the status of cytosine methylation DNA samples. Results A total of 7 708 bands were obtained.Among them 5 636 bands,each representing a recognition site cleaved by one or both of the isoschizomers(Hpa Ⅱ and Msp Ⅰ),were amplified.Furthermore,methylation patterns varied among the four polyploids:heptaploid,octoploid,nonuploid,and decaploid.Total and full methylation levels in P.ternata were 54%-58% and 24.1%-24.3%.All types of MSAP patterns detected in the study belonged to two classes,type Ⅰ and Ⅱ. 52.5% of detected bands belonged to Type Ⅰ; Another 47.5% were type Ⅱ,which showed the methylation differences among the four polyploids. Conclusion The results demonstrate DNA methylation events occur in P.ternata and the general methylation levels are higher.