一、Term
Gel electrophoresis凝胶电泳
agarose gel琼脂糖凝胶（大部分片段核酸）
Polyacrylamide gel聚丙烯酰胺凝胶（小片段、需分辨率高；或蛋白电泳）
Protein electrophoresis蛋白电泳
PAGE: polyacrylamide gel electrophoresis(聚丙烯酰胺凝胶电泳)
SDS-PAGE：SDS-聚丙烯酰胺凝胶电泳
Detergent洗涤剂：SDS十二烷基磺酸钠
Comassie Stain of Protein Gel考马斯亮蓝染色蛋白胶
Two-dimensional gel electrophoresis (双向凝胶电泳)
Isoelectic focusing gel等电聚焦胶（双向凝胶电泳的x轴，y轴用SDS-PAGE）
MALDI-TOF(基质辅助的激光解析电离-飞行 时间质谱)
ESI-MS(电喷雾质谱)
Peptide mass fingerprinting肽质量指纹图谱(PMF)
PCR: Polymerase chain reaction聚合酶链式反应
RT-PCR （reverse transcriptase）逆转录PCR
Q-PCR （real time quantitative PCR）实时定量PCR
cDNA（complementary DNA）：RNA互补DNA，转录本逆转录
cDNA library：cDNA文库，某个样本的所有转录本之和，克隆到载体上，导入大肠杆菌中
Genomic library：基因组文库
DNA library：DNA文库，包括cDNA library和genomic library
The threshold line阈值
The Ct value Ct值，cycle quantification value，到达阈值时的周期数
restriction endonucleases限制性内切酶
cloning vectors克隆载体
Nucleases核酸酶（phosphodiester bond磷酸二酯键）
Phosphatases磷酸酶（phosphomonoester bond单磷酸酯键）
Endonuclease核酸内切酶
Exonuclease核酸外切酶
Isoschizomers (同裂酶)
Isocaudomers (同尾酶)
Recombinant DNA重组DNA
Vectors (载体)
MCS ( Multiple cloning site,多克隆位点)
ORI (复制子)
Phages噬菌体
Replica plating影印培养
pUC Plasmid pUC质粒（MCS有LacZ基因）
β-galactosidase (β-半 乳糖苷酶)
lytic cycle (裂解循环)
lysogenic state(溶源循环)
Cosmids (cos site-carrying plasmid (粘粒、柯斯载体)
ARS (autonomous replicating sequence)自主复制序列
Shuttle vector 穿梭载体
Expression vectors表达载体
Alkaline phosphatase碱性磷酸酶（去除载体5‘端磷酸，防止载体自连）
Klenow fragment：DNApol I大片段，去除5‘-3‘核酸外切酶活性
Restriction Mapping: (内切酶作图)
Inducible expression vector诱导型表达载体
isopropylthiogalactoside (异丙基硫代半乳糖苷, IPTG)
fusion proteins融合蛋白
affinity chromatography (亲和层析法)
Nickel affinity chromatography (镍亲和层析法)
opine冠瘿碱
Binary plasmid (双元质粒)
Helper plasmid (帮助质粒)
Nucleic acid hybridization核酸杂交
Southern blot是用DNA探针检测DNA（看拷贝、基因导入）
Northern blot是用DNA探针检测RNA（看转录本）
Labeled tracers (标记示踪)
Autoradiography放射自显影
Nonradiosctive labeled tracers非放射性示踪
chemiluminescence (化学发光)：大部分northern、southern、westurn都是用这个
densitometer (显像密度计)
Phosphorimaging (磷屏成像)
Satellite (卫星) DNA：short, repeating sequences，CsCl密度梯度离心可以看到
Minisatellite DNA(小卫星DNA): highly polymorphic高度多态. 5-100 bp long, up to 3000 repeats.
Microsatellite DNA ( 微 卫 星 DNA): 2-6 bp sequence repeated to. 50-100 bp lengths, in human genome, about 30000 loci. scattered throughout genome.基因组上位点多
SSR (Simple sequence repeat) =STR (Short Tandem Repeat)= Microsatellite DNA
DNA fingerprinting DNA指纹分析：use a minisatellite DNA as a probe
RFLP(Restriction Fragment Length Polymorphism) 限制性内切酶片段长度多态性
Northern Blot Northern 杂交
Western Blot Western 杂交
In situ DNA hybridization (DNA原位杂交)：定位基因或看有没有特殊的DNA序列
FISH: fluorescence in situ-hybridization (荧光原位杂交)
In situ RNA hybridization (RNA原位杂交)：看基因表达
STR DNA分型技术=SSR DNA分型技术：无需Southern Blot
SNP（single nucleotide polymorphic）单核苷酸多态性
DNA Typing methods for SNP: CAPS, dCAPS
CAPS:Cleaved Amplified Polymorphic Sequence切割的扩增多态性
dCAPS: Derived Cleaved Amplified Polymorphic Sequence衍生的切割的扩增多态性
DNA sequencing DNA测序
Sanger Dideoxy Sequencing双脱氧末端终止法
“Next-generation” sequencing technology二代测序技术
二、判断
1、DNA and RNA molecules are negative charged, they migrate toward to the positive pole
2、The mobilities of fragments are correlated with the log of their molecular weight (or number of base-pairs)
3、要鉴定感兴趣的蛋白具体是哪个蛋白，就用MALDI-TOF跑一个PMF肽质量指纹图谱，与该物种蛋白质的理论数据库进行比对鉴定，简便易行。
4、Purpose of a PCR: make a huge number of copies of a gene.
5、PCR Process：
denaturation 94℃ 1min（双链分开）；
annealing 54℃ 45s（如果要引物特异性高，减少非特异性扩增，就增加退火温度）；
extension 72℃ 2min（聚合酶工作，时间与延伸的长度有关）；
6、不能用PCR的胶中的强度来比较DNA的量，因为PCR扩增是非线性的，并且有平台期，要精确定量，必须用qPCR
7、4-bp cutter cut genome more frequent than 6-bp cutter
8、The first letter is from the genus (属名), the next two letters are from the species (种名). The strain (菌株) designation is included as the forth letter.罗马数字是从这种菌中分离鉴定到的第几种酶
9、利用同尾酶Isocaudomers进行双酶切。
10、Cosmids: engineered λ Phages, can accommodate up to 50kb inserts.Cosmid结合了两者的优点，既有质粒复制的特性又有λ Phages侵染的特性
11、用两个marker来筛选左右臂都连上YAC的
12、DNA转化细胞的方法：热激Chemical transformation、电击Electron transformarion
13、表达载体的两个重要因素：强启动子、5‘非翻译区（核糖体结合区）
14、Advantages of “Eukaryotic proteins made in eukaryotic cells” :
1）Protein will be folded properly
2）proteins are modified in a eukaryotic manner
15、Using a shutter vector (that can replicate in both bacteria and eukaryotic cells), the initial cloning is usually done in E.Coli, then the recombinant DNA is transferred to the eukaryotic cells.
16、Southern 杂交的应用：小卫星DNA做指纹图谱、RFLP
17、Northern Blot：Use DNA as probe、Use to study the expression of a gene
18、Western Blot：蛋白
19、STR DNA分型技术=SSR DNA分型技术
STR (Short Tandem Repeat) DNA分型技术：也叫微卫星序列分析技术或短串联重复序列分析技术，已经成为法医、亲子鉴定领域和DNA数据库建设首选技术。
无需做Southern, 只需对微卫星基因位点进行PCR扩增。
三、简答
1、SDS-PAGE
treat protein molecules with detergent (SDS) to denature (dissociate) the subunits of a protein用洗涤剂(十二烷基硫酸钠)处理蛋白质分子，使蛋白质的亚基变性(解离)
(1)SDS coats all the polypeptides with negative charges, so they all electrophorese toward the anode
十二烷基硫酸钠使所有的多肽都带上负电荷，因此它们都朝阳极
(2)It masks the natural charges of the subunits, so they all electrophorese according to their molecular masses, and not by their native charges.
它掩盖了亚基的天然电荷，因此它们都是根据它们的分子质量而不是它们的天然电荷而进行的.
2、Requirements for a PCR
- Sequence information of the GOI for designing primers
- Appropriate template (DNA)
- Enzymes
- Mg2+ (adjustable可调)
- Buffer (pH and salt concentration盐浓度)
- dNTP (materials for make new DNA molecule)
3、Calculation of Relative Quantification
4、Characteristics of Types II Restriction Endonucleases (RE)
分子克隆常用的II型限制酶：
（1）识别序列和切割序列相同
（2）识别的是回文序列
（3）切割位置固定
（4）产生粘末端或者平末端
5、Essential components of a vector:
1）ORI (复制子): the site where DNA replication begins; to ensure the
replication of the foreign DNA
2）Antibiotic marker gene: to select the host cells with the vector.
3）MCS ( Multiple cloning site,多克隆位点): allow the insertion of foreign DNA
6、Types of vectors
·Plasmids are extra-chromosomal DNA elements, circular dsDNA.
·Phages: l Phages containing linear dsDNA.线性DNA
·Artificial chromosomes contain all the elements that a DNA needs to function as a chromosome in the host organism
1) YAC's are yeast artificial chromosomes
2) BAC's are bacterial artificial chromosomes
7、Phages’ advantages over plasmids:
1. the infection to host cells are much more efficient侵染更高效
2. Accommodate much more foreign DNA(can be used for make library)获得更多外源DNA
8、不同载体的比较
9、酶切作图（！）
10、农杆菌介导
11、双元系统
12、Procedure of a Southern blot
（1）Genomic DNA digestion with restriction endonucleases酶切
（2）Gel electrophoresis of the digested DNA fragments电泳，得到弥散的图（因为是乱切的）
（3）DNA fragments are denatured and transferred into a nitro-cellulose membrane转膜
（4）Using certain labeled probes to hybridize the membrane核酸探针杂交
（5）Wash out the extra probe
（6）Autoradiography or other methods to detect the labeled signal
杂交之前要用非特异性的DNA进行block
13、CAPS
Cleaved Amplified Polymorphic Sequence切割的扩增多态性
