Chapter 1. Methods in molecular biology

Secondary structure
Through biochemical research, people can understand the basic processes and mechanisms of organisms, explore the mechanisms of disease occurrence and development, and provide scientific basis for disease prevention, diagnosis, and treatment.
Biochemistry is an interdisciplinary field that aims to reveal the chemical essence behind life phenomena.
Definition of Biochemistry
Biochemistry helps to increase crop yield and improve quality in agriculture, and contributes to the development of nutritious, safe, and reliable new types of food in the food industry.
Large molecules in living organisms, such as proteins, nucleic acids, and polysaccharides, all have carbon as their core.
Carbon atoms can form complex network structures through covalent bonds, which endow biomolecules with high diversity and stability.

醇)=24:1, mix by inversion and centrifuge at 5000 xg for 10 min at room temperature. • 5. Transfer an aqueous phase to a new tube, add 2/3 volume of isopropanol （异丙醇), mix by gentle inversion and put it into -20 ℃ for at least 1 hour (if necessary). • 6. Centrifuge ca. 5000-10000xg for 10 min, dry pellet and dissolve in TE (5-10 ml).
occurs when DNA is denatured. • 2. Estimation of size: by gel electrophoresis
• 3. Organization: ** Higher plant genomes: Size: 0.2 pg - >100 pg (>500 fold difference) due to repetitive DNA that can account for >80% of some plant genomes. (e.g., maize: 3x109bp/n --- 60-80% repetitive sequence) Single copy: ≥1.5 Kbp in length. Repetitive sequence: 0.15 - 1.0 Kbp in length alternate with single copy sequence.
1% CTAB 0.1% β-mercaptoethanol（巯基乙醇） • ** CTAB: Cethyltrimethylammonium bromide (溴化十六烷基三甲铵 )

“配方”或“方法”
–包括按部就班的操作步骤、试验必需的原材料清单
（原材料包括化学成分、硬件、软件等等）
–包括注释和提醒，提醒使用者在试验过程中需要注意
的问题，以及如何解决问题
–帮助科研人员研究基因行为、疾病治疗新方法、以较
低成本进行新药开发
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What makes a good protocol?
• Complete and thorough instructions • Detailed materials list • Clearly organized • Supporting images/tables • Video Protocols • Trustworthy source
提纲
1
• 什么是Protocols • Springer Protocols • 平台使用介绍
什么是 “protocol”?
2
• Protocol: 实验室指南
• 实验室指南:详细、精确地实验操作记录,主要面向生物化
学、分子生物学、以及生物医学等学科。
–实验室指南是一种标准化的，并可在实验室再现的
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Recreating experiments
• Protocols are used wherever scientific experiments are conducted
• Laboratory methods often first published in research literature –
–Protocol鼻祖亲历亲为(Springerprotocols的主编是全球Protocol标
准的创建人)
–出版500余本广受欢迎及经同行评议的书籍；这些书籍曾在过去20年被

….CTTAAG….
Pst I
Provindencia stuartii 164 Haemophilus influenzae Rd
CTGCAG GACGTC
blunt ends
eg EcoR1 cuts to produce complementary or “sticky” ends. Because it recognizes and cuts this sequence of 6 (6 cutter) , It makes med. -sized pieces. 4-cutter, small and 8 cutter, large pieces.
Methods in Molecular biology
QiuZheng 邱郑 qiuzheng754@
Charpter 4 Molecular Cloning Methods
• Why to clone genes? • How to clone genes in bacteria and in eukaryotes?
• Enzymes that recognize identical sequences are called isoschizomers, although the cutting sits within these sequences are different.
SmaI 5’….CCC GGG.….3’ ….GGG CCC….
Insertional inactivation
pBR322 cloning
• Plasmid and insert cut with Pst I • Join sticky ends • Transform bacteria • Screen for tetracycline resistance and ampicillinsensitive • Foreign DNA interrupts Amp gene