斯坦福大学分子生物学讲义(英文)
分子生物学讲座(8)
四、限制性核酸内切酶的作用
1、切割方式
大部分限制性核酸内切酶识别DNA序列具 有回文结构特征，切断的双链DNA都产生5’ 磷酸基和3’羟基末端。不同限制性核酸内 切酶识别和切割的特异性不同。
5`…GAATTC…3` 3`…CTTAAG…5`
5`…AAGCTT…3` 3`…TTCGAA…5`
EcoR I
一般认为生物技术包括基因工程、细胞工程、 酶工程和发酵工程几方面的内容。基因工程是生 物技术的核心和关键，是主导技术；细胞技术是 生物技术的基础；酶工程是生物技术的条件；发 酵工程是生物技术获得最终产品的手段，四个方 面相互联系的。生物技术是一个综合技术体系， 其中基因工程和细胞融合技术y gene)是编码能与操纵子序 列结合的调控蛋白的基因。调控方式有负调控 (negative regulation)和正调控(positive regulation)。
某些特定的物质能与调控蛋白结合，使调控蛋 白的空间构像发生变化，从而改变其对基因转录 的影响，这些特定物质中凡能引起诱导发生的分 子称为诱导剂(inducer)，能导致阻遏发生的分子 称为阻遏剂或辅助阻遏剂(corepressor) 。
三、限制性核酸内切酶的分类
按限制酶的组成、与修饰酶活性关系，切断 核酸的情况不同，分为三类：
Ⅰ类限制性核酸内切酶：由3种不同亚基构成， 具有修饰酶活性和内切酶活性，它能识别和结合 于特定位点，随机切断识别位点以外的DNA序 列，通常在识别位点周围400-700bp（10005000bp）。这类酶的作用需要Mg2+，S腺苷甲 硫氨酸及ATP。
谢谢大家！
X-gal(5-溴-4-氯-3-吲哚-β-半乳糖苷)也是一 种人工化学合成的半乳糖苷，可被β-半乳糖苷 酶水解产生兰色化合物，因此可以用作β-半乳 糖苷酶活性的指示剂。IPTG和X-gal被广泛应 用在基因工程的工作中。
分子生物学英文原版
分子生物学英文原版Molecular Biology English Original is an essential reference book for students and researchers who areinterested in the study of molecular biology. It provides a detailed exploration of molecular biology research and discoveries, and serves as a comprehensive guide to different techniques and methodologies used in the field. In this article, we will discuss the book in detail, step by step.Step 1: Overview of the BookThe book starts with an introduction to molecular biology and an overview of the key concepts and principles involved. It provides a history of molecular biology research and the key discoveries that led to the development of the field. The book then goes on to cover different topics such as DNA replication, transcription, translation, gene regulation, and genetic engineering.Step 2: Understanding the ConceptsThe book explains the concepts and theories of molecular biology in a clear and concise manner. It provides detailed explanations of different techniques and methodologies usedin molecular biology research, including DNA sequencing, PCR, gel electrophoresis, gene cloning, and protein purification. It also includes detailed walkthroughs of various experiments and provides step-by-step procedures for performing them.Step 3: Real-life ApplicationsThe book provides real-life applications of molecular biology research and technologies. It describes how molecular biology has revolutionized medical science, plant science,and bioengineering. It discusses the use of molecular biology techniques in disease diagnosis, gene therapy, and drug development. It also talks about how molecular biology isused to engineer crops with desirable traits and to develop new pharmaceuticals.Step 4: Future DevelopmentsThe book ends with a discussion of future developmentsin molecular biology research. It highlights the potentialuse of CRISPR-Cas9 gene editing technology, which has revolutionized gene editing and holds great promise for the treatment of genetic diseases. It also discusses the use of molecular biology in the development of personalized medicine, where treatments are tailored to an individual's genetic makeup.In Conclusion, Molecular Biology English Original is an indispensable resource for students and researchers in thefield of molecular biology. It provides a comprehensive overview of molecular biology research and discoveries, and serves as a guide to different techniques and methodologies used in the field. The book is an excellent reference toolfor those interested in the real-world applications of molecular biology and its future developments.。
分子生物学-3-chapter 21
Useful features of the flies in
experimental research :
Fecundity
Rapid life cycle
Four chromosomes (two large autosomes, a smaller X, and a very small fourth chromosome)
given physical segments of chromosomes , (phenotypes of flies white eyes were correlated
with deletions in the chromosomes
Bridges used polytene chormosomes to determine a physical map of the
Polytene chromosomes
In the salivary gland, extensive endoreplication without mitosis,
Giant chromosomes composed of approximately 1000 copies of each chromatid
Embryos that contain two copies of the
balancer chromosome die, because of the invresions produce recessive disruption in critical genes. In addition, embryos that contain two copies of “nomal” chromosome die, because they are homozygous for
【免费下载】分子生物学讲义
2 DNA与染色体2.1 DNA的一般性质2.2 DNA的二级结构及多态性2.3 DNA的超螺旋结构2.4 染色体结构2.1 DNA的一般性质1868年，瑞士的内科医生Friedrich Miescher从外科医院包扎伤口的绷带上的脓细胞核中提取到一种富含磷元素的酸性化合物，将其称为核质(nuclein)；后来他又从鲑鱼精子中分离出类似的物质，并指出它是由一种碱性蛋白质与一种酸性物质组成的。
20年后称为核酸(nucleic acids)，其功能不清楚。
1944年Avery等，发现从S型肺炎球菌中提取的DNA与R型肺炎球菌混合后，能使某些R型菌转化为S型菌，且转化率与DNA纯度呈正相关。
若将DNA预先用DNA酶降解，转化就不发生。
结论是：S型菌的DNA将其遗传特性传给了R型菌，DNA就是遗传物质。
1952年A.D.Hershey和M.Cha-se用35S和32P双标记T2噬菌体证明DNA是遗传物质。
噬菌体在35S培养基中外壳蛋白被标记；在32P培养基中DNA被标记；这种双标记的噬菌体感染大肠杆菌时，DNA进入细胞大量复制（只有亲本DNA链才有32P）并装配成子代颗粒，只有少量的噬菌体有32P，而无外壳有35S。
2.1.1 多核苷酸链2.1.1.1 核酸的化学成分核酸是生物体内的高分子化合物。
它包括脱氧核糖核酸(deoxyribonucleic acid，DNA)和核糖核酸(ribonucleic acid，RNA)两大类。
DNA和RNA是由单个核苷酸(nucleotide)头尾相连而形成的。
RNA平均长度大约为2000个核苷酸；人DNA可长达3×109个核苷酸。
核苷酸是由碱基、戊糖和磷酸构成。
碱基(base)：嘌呤(purine) ：腺嘌呤(adenine，A)、鸟嘌呤(guanine，G)；嘧啶(pyrimi-dine) ：胞嘧啶(cytosine，C)、胸腺嘧啶（thymine，T)、尿嘧啶(uracil，U)。
英汉对照分子生物学导论(王勇)Chapter_0
3. How to learn?
Because we have so many things to learn, it is very easy to get lost in these details, and so before we begin to study, we need some guiding principles to give a sense of order to many of these details. 因为我们有这么多内 容要学，而一旦深入 到细节中又很容易迷 失方向，因此在开始 学习之前，我们需要 一些指导原则来帮助 梳理这些细节内容。
Benjamin Lewin Gene VIII
4 / 20
Introduction / 引 言
1. What is molecular biology? 什么是分子生物学？
2. What should we learn? 我们应该学习什么内容？
3. How to learn? 如何学习？
5 / 20
19 / 20
A wonderful molecular world !
20 / 20
1. What is molecular biology?
Molecular biology: “How does a set of molecules make a dynamic living organism?” The most fundamental feature of all known life is the ability to reproduce and the ability to grow. If organisms were not able to reproduce, life would be impossible.
分子生物学(全套课件557P)
分子生物学（全套课件557P）简介分子生物学是研究生物分子结构、功能和相互作用的学科。
它涉及到核酸、蛋白质和其他生物分子的研究，以及它们在细胞和生物体中的功能。
本文档是一套全面的分子生物学课件，共有557页。
本课件旨在帮助读者系统地了解分子生物学的各个方面，包括基本的分子生物学原理、实验技术、研究方法以及应用等。
目录1.第一章：分子生物学概述2.第二章：DNA结构与功能3.第三章：RNA结构与功能4.第四章：蛋白质结构与功能5.第五章：基因表达调控6.第六章：基因突变与遗传变异7.第七章：分子生物学实验技术8.第八章：分子生物学研究方法9.第九章：分子生物学的应用领域第一章：分子生物学概述1.1 什么是分子生物学分子生物学是研究生物体内分子的结构、功能以及相互作用的学科。
它涉及到DNA、RNA、蛋白质等生物分子的研究，以及它们在细胞和生物体中的功能。
1.2 分子生物学的历史与发展分子生物学起源于20世纪50年代，当时发现DNA是物质遗传信息的携带者后，科学家们开始研究DNA的结构和功能，从而奠定了现代分子生物学的基础。
1.3 分子生物学的重要性分子生物学的研究对于了解生命的本质和机理至关重要。
它不仅有助于解释遗传现象，还可以揭示细胞的结构、功能和调控机制，甚至为疾病的诊断和治疗提供理论基础。
2.1 DNA的组成与结构DNA是由基因序列组成的生物分子，它由核苷酸组成。
本节将介绍DNA的基本结构、双螺旋结构和碱基对的配对方式。
2.2 DNA复制与遗传信息传递DNA复制是细胞分裂过程中最重要的事件之一，它确保了遗传信息的传递和稳定性。
本节将介绍DNA复制的过程和机制。
2.3 DNA修复与突变DNA在生物体内容易受到各种外界因素的损伤，因此细胞拥有多种修复机制来修复DNA损伤。
本节将介绍DNA修复的方式和维护基因组稳定性的重要性。
3.1 RNA的种类与功能RNA是DNA转录的产物，它在细胞内发挥着多种功能，包括mRNA的编码信息传递、tRNA的氨基酸运载和rRNA的构建核糖体等。
哈佛大学分子生物学讲义Replication3A
late G1
MCM
S
MCM
"Pre-Replicative Complex"
early G1 M G2
免疫学信息网
ORC
Eukaryotic initiation complex
ORC : A six subunit protein complex which has been implicated as being the eukaryotic DNA replication initiator protein. Subunits are named according to their size, with ORC1 being the largest and ORC6 being the smallest subunit. Yeast ORC specifically binds to replication origins in an ATP dependent manner and has been shown to possess ATPase activity. CDC6/Cdc18 : An essential factor for the assembly of the prereplicative complexes that co-operates with Cdt1 to load MCM2-7 Proteolyzed in yeasts or exported out of the nucleus in mammalian cells at the G1- S transition. Overexpression of Cdc6 in yeast causes multiple rounds of DNA replication without intervening mitosis, making it a critical regulator of DNA replication. MCM2-7 : A family of six related proteins (MCM2-MCM7) which seem to function together in a large multi-subunit protein complex. The role of MCMís in DNA replication is still speculative, ranging from mediation of protein-DNA interactions to DNA helicase activity.
(完整word版)《分子生物学》教案
三、原核生物染色体和基因(50＇)
四、小结（5＇）
三、授课重点
1.以大肠杆菌为代表的原核生物基因结构的组织特点。
2.C值及C值矛盾,以及如何根据Cot1/2 值来确定基因组的复杂性
2、C值及C值矛盾,以及如何根据Cot1/2 值来确定基因组的复杂性
四、授课难点
1.E。coli的基因结构的特点
二、嘧啶二聚体的产生（10＇）
三、二聚体修复的机制(60＇)
四、限制和修饰（10＇）
五、小结(5＇）
三、授课重点
损伤修复的几种形式及其修复过程
四、授课难点
1.核苷酸切除修复和重组修复的修复时期及简单过程
2.SOS修复系统
2、SOS修复系统
五、授课方法与课前准备
损伤修复的几种形式及其修复过程式本节的重点, 为了更好的说明各个修复过程,在网上下载了大量的图片。
为了学生更好的理解DNA和RNA在结构上的区别, 加入了大量的图片。
制作PPT教学课件、讲授、结合板书
六、教研室审查意见
主任签字
一、题 目
第二章DNA的结构
第三节DNA的二级结构
第四节 DNA的物理化学性质
二、单元教学目标与课时分配（共100min)
一、上次课内容回顾（10＇）
二、DNA的二级结构（50＇）
《Gene Ⅶ》 Lewin.B。 Oxford University Press 1999
一、题 目
第一章绪论
第一节 分子生物学概念
第二节 分子生物学的发展历程
二、单元教学目标与课时分配（共100min）
一、分子生物学概念(50＇)
二、分子生物学的发展简史（45＇)
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Top view with template-primer: Polymerase site And proofreading site
Topoisomerases relax DNA by changing the DNA linking number
Looping the lagging strand to make both polymerases move in the same direction
The discovery of DNA polymerase.
Arthur Kornberg and Bob Lehman pursued an enzyme in bacterial extracts that would elongate a chain of deoxyribonucleic acid just like glycogen synthase elongates a chain of glycogen.
Wait a minute!
John Cairns mutated the gene for DNA polymerase, polA, and the bacteria grew just fine!
Either the polymerase hypothesis was all wrong,…… or there were other DNA polymerases in E. coli that carried out DNA synthesis in the polA strains.
* Topoisomerases I change the linking number in steps of 1. They pass a single DNA strand through a nick.Topoisomerase I is a protein of the metaphase chromosome scaffold. * In interphase, topoisomerase is bound to the nuclear matrix. * The DNA replication machinery also appears bound to the matrix. * Inhibitor (camptothecin) also used in chemotherapy.
130 dnaE |
5'-3' polymerase
27.5 dnaQ | POL III CORE 5'-3'
(mutD)
exonuc leas e
10
|
3'-5'
exonuc leas e
71 dnaX
47.5 dnaX |
35
|
' 33
| COMPLEX
15
|
12
|
ATP dependent clampl oade r
Which polymerase is processive?
P
POL
dNTP
Challenge with vast excess of cold primer-template
Gel electrophoresis of products
Challenge - +
-+
POL-X
POL-Y
POLIII, subunit
40.6 dnaN CLAMP
processivity
factor
DNA POLYMERASE III
Sub Gene Bacterial unit
Function
Eukaryotic
dnaE |
dnaQ | POL III
(mutD) CORE
|
5'-3' polyme rase 3'-5' exonuc leas e 5'-3' exonuc leas e
PCNA
Clamp loaders hydrolyze ATP to load clamp
Clamp-loader ATP
ATP
How does one prove that the clamp ring is opened during loading?
Clamp
ATP
ADP + PPi
3慜H
Structure of a DNA polymerase (gp43 from phage RB69)
* Topoisomerases II change the linking number in steps of 2 by passing both strands of double-stranded DNA through a break. * Eukaryotic topoisomerases isolated to date only relax supercoiled DNA, while prokaryotic topoisomerases (gyrases) can, given ATP, add supercoils. * TopoII releases catenated daughter molecules at the end of replication. Inhibitors like etoposide are used in chemotherapy.
DNA POL DNA POL Fen1
dnaX
dnaX |
|
ATP
'
|
dependent RF-C
COMPLEX clamploader
|
|
dnaN CLAMP processivity PCNA factor
CONSERVATION FROM PROKARYOTES TO EUKARYOTES
The enzymatic activity was unusual:
1) Needed a template which dictates what nucleotide was added: substrate was directing enzymatic activity 2) Needed a primer annealed to the template.
3H Thymidine incorporation (pmol) Phosphate (M)
I
600 200
III
100
+ NEM
polA + (wild type) 0.4M II
0.2M
200
polA- (Cairns) II
0.4M
III
100
0.2MI2030 Nhomakorabea40
Fractions
Sub kDa Gene Subassembly unit