ROLE OF CYTOSINE METHYLATION IN C. CINEREUS

胞嘧啶甲基化在华支睾吸虫中的作用 已知CpG甲基化参与了基因沉默和 负责基因组印记。我们希望了解一下 胞嘧啶甲基化在真菌系统中的后果 鸡腿菇(Coprinus Cinereus)在这项研究中，我们确定了位置和 色氨酸合成酶中5-甲基胞嘧啶残基的含量 基因(TRPL)。以前的研究表明，胞嘧啶甲基化 可能是由于C。 灰熊基因组。而且，我们利用甲基化酶来 外源基因潮霉素B中的甲基化胞嘧啶残基 磷酸转移酶(PHT)。我们使用基因组测序方案 由此胞嘧啶在治疗时被脱氨基为尿嘧啶 亚硫酸氢钠保持不变，5-甲基胞嘧啶保持不变。 因此，5-甲基胞嘧啶(M5C)残基的存在是 由序列数据中剩余的胞嘧啶指示。我们 将分析六个同时具有这两种基因的四分体的TRPL基因序列 Trp+和Trp-子代直接比较不同NCEs 沉默和非沉默基因之间的甲基化。这 序列分析将确定是否存在位点特异性 胞嘧啶甲基化，如果这种甲基化只发生在 在CpG二核苷酸。此外，我们可能会了解到，如果具体 甲基化胞嘧啶是胞嘧啶的必要条件和充分条件 使TRPL基因失活。我们将进一步确定是否 当外源基因是 在体外甲基化，并转化到华南星。C。 Cinereus已被证明是研究甲基化的模型系统 并可能给我们提供关于基因沉默是如何发生在 物种，可能作为一个整体存在于真核生物中。

Examining the Role of Cytosine Methylation in C. cinereus CpG methylation is known to be involved for gene silencing and to be responsible for genomic imprinting. We wish to understand the consequences of cytosine methylation in the fungal system Coprinus cinereus. In this study we identify the location and extent of 5-methyl cytosine residues in the tryptophan synthetase gene (trpl). It was shown previously that cytosine methylation can occur as a result of a duplication of the trpl gene in the C. cinereus genome. And, we make use of methylating enzymes to methylate cytosine residues in the exogenous gene Hygromycin B Phosphotransferase (PHT). We use a genomic sequencing protocol whereby cytosine is deaminated to uracil upon treatment with sodium bisulfite while 5-methyl cytosine remains unchanged. Thus, the presence of 5-methyl cytosine (m5C) residues is indicated by the remaining cytosines in the sequence data. We will analyze the trpl gene sequence in six tetrads that have both Trp+ and Trp- progeny to make a direct comparison of differe nces in methylation between silenced and unsilenced genes. This sequence analysis will determine if there is site specificity for cytosine methylation, and if this methylation occurs exclusively at CpG dinucleotides. In addition, we may learn if specific methylated cytosines are cytosines are necessary and sufficient to render the trpl gene inactive. We will further determine if maintenance methylation exists when an exogenous gene is methylated in vitro and transformed into C. cinereus. C. cinereus has proven to be a model system for studying methylation and may give us clues about how gene silencing occurs in this species and possibly in eukaryotes as a whole.