| 其它PCR方法 | | 点击： 作者： 来源： 时间： 2006-11-06 本站论坛 |
|  |
· (Molecular Biology Techniques Manual)
The followings are described in detail
Recommended Reagent Concentrations
Recommended Reaction Conditions
'Hot Start' PCR
Asymmetric PCR for ssDNA Production
Detecting Products
Labeling PCR Products with Digoxigenin
Cleaning PCR Products
Sequencing PCR Products
Cloning PCR Products
- (BMB)
This manual contains detailed protocols on performing PCR as well as preparation of templates and post-PCR clean-up. Important applications such as PRINS, cycle sequencing, detection of human chromosomes, and non-radioactive labeling are discussed.
Core Sample PCR
· (NWFSC)
A method to re-PCR unique bands from products of mixed size
· (FMC)
Touch Down PCR
· (Alkami Biosystems)
Hot-Start PCR
· (Molecular Biology Techniques Manual)
Asymmetric PCR
· (Molecular Biology Techniques Manual)
Inverse PCR
· (E. Jay Rehm,
Berkeley Drosophila Genome Project)
· (PMCI)
To Isolate DNA Adjacent To Known Sequence in Genomic DNA
· (Gottsschling Lab)
Degenerate PCR
· (Molecular Biology Techniques Manual)
For amplification of cognate sequences from different organisms, or for 'evolutionary PCR', one may increase the chances of getting product by designing 'degenerate' primers: these would in fact be a set of primers which have a number of options at several positions in the sequence so as to allow annealing to and amplification of a usr/localiety of related sequences.
· (Michael Koelle)
· (Michael Koelle) (at another server)
Detailed guide in degenerate PCR, including primer design, reaction conditioning and product analysis.
· (Atle M. Bones)
PCR-Elisa
· (NUNC)
Run PCR with internal amplification control. After completion of the competitive PCR the target DNA and the control DNA are detected by colorimetric reaction in separate wells on the microtiter plate.
Others
- (Gary A. J. Brown)
Amplify DNA from a drop of Blood.
· (Schneitz Lab)
Use tissue directly for PCR without DNA extraction
· PCR DNA Biotinylation (KPL)
A rapid method for biotinylating DNA probes through incorporation of biotin-N4-dCTP via a thermostable DNA polymerase in the polymerase chain reaction
· ( Cindy Troxell)
Colony PCR can be used to identify colonies where your favorite gene (yfg) has been replaced with a marker gene by homologous recombination, and to distinguish homologous recombination events from non-homologous. It can also be used to identify colonies from a tetrad that carry a particular gene replacement. This is not a substitute for Southern blotting, but an adjunct.
· (D. Amberg)
It works either for E coli and yeast
· (Namjin Chung)
Zymolyase Method for screening yeast transformants
· (Detection of Immobilized Amplified Products) (NUNC)
Introduction to DIAPOPS
· (NUNC)
· (NWFSC)
Use of PCR in site-directed mutagenesis accomplishes strand separation by using a denaturing step to separate the complementing strands and allowing efficient polymerization of the PCR primers. PCR site-directed methods thus allow site-specific mutations to be incorporated in virtually any double-stranded plasmid; eliminating the need for M13-based vectors or single-stranded rescue.
· (Molecular Biology Techniques Manual)
How to calculate primer and nucleotide concentration
· (Amersham)
Guidelines for preparation of radioactively-labeled PCR products for use as probes
· (Donis-Keller lab)
This method is used to sequence directly a specific PCR product after large scale amplification. The reaction should be carried out in the usual fashion, except that after optimization of annealing temperature and other conditions in the 5 µl volume, a large scale 50 µl (10X) total reaction volume should be used to generate sufficient PCR product for sequencing.
· (Genetics Resources CORE Facility at Johns Hopkins U)
Detailed method for preparing PCR products for direct sequencing
- 96 Well Plate Purification of PCR products
Purification with Sepahcryl S-300 or S-500 - in filter bottom plates 96-well silent screen plate, Purification with Guanidine-HCl in MES buffer - in glass filter plates
Purification with Guanidine-HCl in KAc buffer
- Protocol for PCR of M13
To generate second strands for reverse sequencing of M13 templates
- (E. Beasley and D. Amberg)
- (Jeffrey D. Newman)
- (Linda Riles)
The selectable marker for deletion (HIS3) is amplified by PCR using oligos that contain homology to the HIS3 gene and also 45 bp of homology immediately 5' and 3' to the gene being deleted. The PCR product is used to transform yeast to His+, which removes the gene from START to STOP. Correct gene deletion is verified by PCR of yeast colonies, using a universal test oligo in HIS3, and an oligo in flanking chromosomal sequence. Only correctly deleted transformants yield a PCR product.
· (Saccharomyces Genome Deletion Project)
A colony PCR strategy is used to identify successful deletions. This method is fast, reliable, and amenable to a 96-well format
· ()
PCR in teacup can simply be defined as PCR without a PCR machine. This 'old-fashioned' method uses hot plates and water baths to go through the amplification process. Because of its tediousness it is not the most successful way to perform a polymerase chain reaction, but its advantages are that it is inexpensive and simple.
· (Jackson Labs)
Using tail tissue directly for PCR after proteinase K digestion.
· (Jackson Labs)
Simple method for preparing DNA from blood for PCR without organic extraction
· attB Adapter PCR Protocol (LTI)
The Adapter PCR protocol allows for shorter primers to amplify attB-PCR products by utilizing four primers instead of the usual two in a PCR reaction. This protocol is particularly useful when cloning large numbers of genes (universal attB adapter primers can be synthesized once and used hundreds of time), or when additional sequences are added to the template specific primer, such as a TEV protease cleavage site, which result in a long primer.
· (Stanford Human Genome Center)
A protocol used to assay for the presence of an STS for radiation hybrid mapping
上一篇：原位PCR 下一篇：反向PCR | | |
|
|
|
