PCR技术

Degenerate PCR is in most respects identical to ordinary PCR but with one major difference. Instead of using specific PCR primers with a given sequence you use mixed PCR primers. That is if you do not ...

Isolate partially purified chromosomal DNA using a modification of the Magic MiniPreps DNA Purification System from Promega (see atta ched) as follows. - Spin down 500 μl of an overnight culture in a ...

Following PCR you often want to get rid of the PCR primers and Taq polymerase before the next step. This is necessary for sequencing PCR products or TA cloning. There is no clean up needed following g ...

Multiplex_protocol_NP2006.pdf

1) Immunocapture stageCoating buffer: 15 mM Na2CO3; 35 mM NaHCO3 and 3 mM NaN3 per liter (pH 9.6). Extraction buffer: (20 mM Tris-HCL (pH 8.0) 138 mM NaCl 1 mM PVP 0.05% Tween-20 3 mM KCl and 3 mM Na ...

DescriptionSingle primer PCR allows amplification from known to unknown regions in chromosomes phage plasmids large PCR products and other sources of DNA.At sufficiently low stringency any primer will ...

Colony PCR is useful in determining whether or not a specific colony on a plate has a sequence you desire. Primers for the specific sequence should be used when preparing the reaction cocktail. Exampl ...

TAIL is a series of reactions that are intended to map where a T-DNA (transfer DNA) has inserted within the genome. The main components of the 3 reactions are the AD (Arbitrary Degenerate) primers bor ...

RT-PCR or reverse transcription PCR refers to PCR that uses product of an RT reaction as template. In effect the PCR amplifies cDNA fragments. In one-tube RT-PCR RNA and PCR primers are added to a rea ...

RNA Extraction from Histologic SectionsUnstained 4 µm thick sections of formalin fixed paraffin embedded liver biopsies were transferred from glass slides to 1.5 ml non-siliconized microcentrifuge tub ...

Cut PVDF membrane to the appropriate size activate with absolute methanol for 5 sec and incubate in distilled water for 5 min. For electroblotting equilibrate in transfer buffer and follow the standar ...

The RT-PCR method can be used not only to detect specific mRNAs but also to semi-quantitate their levels. Thus one can compare levels of transcripts in different samples. This can be done in two diffe ...

SummaryQuantitative PCR is a method used to detect relative or absolute gene expression level. All qPCR involves the use of fluorescence to detect the threshold cycle (Ct) during PCR when the level of ...

Why PrimerBank?PrimerBank is a public resource for PCR primers. These primers are designed for gene expression detection or quantification (real-time PCR). There are several ways to search for primers ...

Quantitative real time PCR (QRT-PCR) has become one of the most widely used methods of assessing gene expression owing to its sensitivity and reduced run time compared with traditional methods such as ...

Adapted from TaqMan® One-Step RT-PCR Master Mix Reagents Kit InstructionDesign of Probes Keep the G-C content in the 20 to 80% range. Avoid runs of an identical nucleotide. This is especially true fo ...

Steps for Standard PCR ReactionDesign primers. In general primers should have the following properties: Length of 18-24 bases 40-60% G/C content Start and end with 1-2 G/C pairs Melting temperature (T ...

Comparison of normalisation methods There is an ongoing debate what is the best way to normalise qPCR data. Reference genes are the most common method although single unverified reference genes invali ...

利用实时定量PCR和2－△△CT法分析基因相对表达量　　 METHODS 25 402�408 (2001)Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2－△△CT MethodKenneth J. Livaundefined and Thomas D. Schmittgen¶undefinedAppli ...

RT-PCR PROTOCOL材料与方法………………………………………………………… 1．材料 ………………………………………………………1.1 供试用组织(细胞)…………………………………1.2 主要仪器设备………………………………………1.3 主要试剂……………………………………………1.4 工具酶及试剂盒…………………………………… 2.实验方法…………………………………………… ...