1、宁波大学宁波大学 2015 年攻读年攻读博博士学位研究生士学位研究生 入入 学学 考考 试试 试试 题题(B 卷卷) (答案必须写在答题纸上) 考试科目考试科目: 生物化学与分子生物学生物化学与分子生物学 科目代码：科目代码： 2607 适用专业适用专业: 水产养殖水产养殖/渔业资源渔业资源/水产资源综合利用水产资源综合利用 第 1 页 共 2 页 一、名词解释一、名词解释 （共（共 40 分，每分，每小小题题 4 分）分） 1. Open reading frame(ORF) 2. Reverse transcriptase 3. Edman degradation 4. RNA inter
2、ference 5. Protein chip technology 6. Yeast two-hybrid system 7. Respiratory chain 8. Two-dimensional electrophoresis 9. Allosteric effect of protein 10. Glycolysis and gluconeogenesis 二、二、问答题（共问答题（共 60 分）分） 1. 什么是生物学中心法则？（6 分） 2. 什么是 Southern 印迹？简述该实验的原理。 （6 分） 3. 真核生物基因转录水平的调控机制是什么？（8 分） 4. 简述乙酰 C
3、oA 在动物体内的来源及其去路。 （8 分） 5. 胞内酶分离纯化须通过哪些步骤？应注意哪些问题？并说明利用葡聚糖凝胶过滤层析测定该酶蛋白分子量的主要操作步骤。 （12 分） 6. 请对文章摘要中划线的名词进行解释：RACE technique，phylogenetic tree，introns and exons， real-time PCR，并根据摘要内容结合所学知识写出得到重组 TNFalpha 蛋白的详细步骤。 （20分） 宁波大学宁波大学 2015 年攻读年攻读博博士学位研究生士学位研究生 入入 学学 考考 试试 试试 题题(B 卷卷) (答案必须写在答题纸上) 考试科目考试科目:
4、生物化学与分子生物学生物化学与分子生物学 科目代码：科目代码： 2607 适用专业适用专业: 水产养殖水产养殖/渔业资源渔业资源/水产资源综合利用水产资源综合利用 第 2 页 共 2 页 The tumor necrosis factor (TNF) superfamily is composed by several proteins with similar structure and functions. Using the RACE technique, we have cloned and sequenced the turbot TNF cDNA. The analysis of
5、its sequence showed several conserved motifs characteristic of members of the TNFalpha family. A phylogenetic tree constructed with different TNFs of fish and mammals grouped our sequence within the fish TNFalpha cluster. Therefore, the turbot TNF here studied was identified as TNFalpha. The complet
6、e TNFalpha gene was obtained by gene walking, and, similarly to the other known fish TNFalpha genes, presented three introns and four exons. A real-time PCR was designed to study the turbot TNFalpha expression in vivo using as stimulus the bacteria Vibrio pelagius strain Hq222. The expression of the
7、 cytokine happened early after injection. In addition, TNFalpha expression was in general higher in kidney than in liver, as expected since the former is the haematopoietic organ of fish. The turbot recombinant TNFalpha (rTNFalpha) was obtained by IPTG induction of bacteria transformed with the pET1
8、5b-TNFalpha construct, and it was purified in native conditions. The recombinant protein was approximately 20 kDa in size, and its biological activity was assessed in vitro. No effect of the rTNFalpha neither alone nor in combination with LPS was observed on the chemiluminescence activity of turbot macrophages at any time tested.
