我最近也在做msp，什么结果也没有。有空多交流qq：110137188

zitong1983您好，非常感谢您对NEB的信任和支持。关于甲基化酶，包装中提供的3个EP管分别是，CpG甲基转移酶、10*NEBuffer 2以及SAM（S-腺苷甲硫氨酸）。在使用中您可以看每支EP管上的标签进行区分。以下是recommended protocol for methylation of genomic DNA,请参考：Dilute SAM to 1600 μM using the supplied 32 mM stock. a. SAM 32 mM stock 1 μl b. Nuclease free water 19 μlAdd in order: a. Nuclease free water 14 μl b. 10X NEBuffer 2 2 μl c. Diluted SAM from step 1 2 μl d. Genomic DNA (1 μg) 1 μl e. SssI methylase (4 U/μl) 1 μlMix, Pipette up and down at least six times.Incubate one hour at 37°C.Stop the reaction by heating at 65°C for 20 minutes. Notes: 1. The volume of DNA can be increased to 5 μl. When using more dilute DNA increase the reaction volume to 50 μl. Using too much DNA volume in the reaction can cause inhibition by changing the pH or salt concentration of the reaction. 2. Up to 4 μg of DNA can be methylated in a 20 μl reaction. The SAM concentration should be adjusted to 640 μM. Concentrated SssI M0226M (1 μl of 20,000 U/ml) should be used. 3. The incubation time can be increased to 4 hours. Overnight incubations do not give significant increases in methylation. 4. The above protocol can also be used for other types of DNA including plasmids and purified PCR products. Example of a large-scale methylation: 1. Add in order: a. Nuclease free water 220 μl b. 10X NEBuffer 2 50 μl c. SAM 32 mM 10 μl d. Lambda DNA (500 μg/ml 200 μl e. SssI methylase (20 U/μl) 20 μl 2. Mix, Pipette up and down at least six times. 3. Incubate 2 hours at 37°C. 4. Stop the reaction by heating at 65°C for 20 minutes. The DNA can be purified by phenol extraction followed by ethanol precipitation or by using a commercial DNA purification kit. For long-term storage at -20°C, suspend in TE.您也可以登录NEB的英文网站，进行查阅。在使用过程中，建议您也要看一下说明书中提到的注意事项，并根据您的底物来调整甲基供体SAM的浓度用量。同时，SAM的保质期相对较短，大概六个月，所以如果您购买产品后并没有马上使用，那在使用前一定要注意看一下SAM的效期。祝您实验顺利，有任何问题欢迎随时讨论。谢谢！

NEB，您好！我最近用了NEB公司的SssI甲基转移酶，就是按照你提供的protocol来做的。但是当我用HpaII内切酶去切的时候，仍然能切开。有哪些原因呢？还有什么方法可以证明我的质粒被甲基化了呢？谢谢回复！