PCR Protocol--PCR Elisa

What is PCR Elisa?
PCR products are labeled (e.g., by digoxigenin) during amplification. A capture probe specific to the PCR amplicon is used to immobilize the amplicon to well plate. ELISA against the label (e.g., anti-digoxigenin) is then used to quantitate PCR products.

PCR-ELISA, an alternative to real time PCR for detecting.
PCR-ELISA is an alternative to real time PCR used to detect sequences within the polymerase chain reaction products.www.btc-bti.com/pcrelisa.htm

Mycoplasma Detection by the Mycoplasma PCR ELISAMycoplasma Detection by the. Mycoplasma PCR ELISA. Introduction. The most frequent contaminants of animal ...www.roche-applied-science.com/PROD_INF/BIOCHEMI/jlp33-35.pdf B

Telomerase PCR ELISATelomerase PCR ELISA. For fast and sensitive detection of ... The indicated number of cells were analyzed in the Telomerase PCR ELISA.Assays were ...www.roche-applied-science.com/PROD_INF/BIOCHEMI/No.4_96/p7-8.pdf

Optimisation of one-tube PCR Elisa to detect femtogram amounts of genomic DNA.
A simple, high-throughput, low-cost polymerase chain reaction-enzyme linked immunosorbent assay (PCR-ELISA) protocol that detects the presence of 4 fg of ...linkinghub.elsevier.com/retrieve/pii/S0167701202000556

Qualitative PCR-ELISA protocol for the detection and typing of viral genomes.
Nature Protocols is an interactive online resource for all laboratory protocols relevant to biological and biomedical research. Nature Protocols includes a ...npg.nature.com/nprot/journal/v2/n10/abs/nprot.2007.311.html

PCR Protocol--Nested PCR

Nested PCR

What is nested PCR? Nested PCR is a variation of the polymerase chain reaction (PCR), in that two pairs (instead of one pair) of PCR primers are used to amplify a fragment.

The first pair of PCR primers amplify a fragment similar to a standard PCR. However, a second pair of primers called nested primers (as they lie / are nested within the first fragment) bind inside the first PCR product fragment to allow amplification of a second PCR product which is shorter than the first one.
The advantage of nested PCR is that if the wrong PCR fragment was amplified, the probability is quite low that the region would be amplified a second time by the second set of primers. Thus, Nested PCR is a very specific PCR amplification.

PCR amplification of cDNA segments by 2 stage nested PCRprotocolNCBI
Standard Nested PCR ProtocolC. elegans Gene Knockout Project, Vancouver Lab
Poison Primer Nested PCR Protocol
Moerman Lab.
Nested PCR and Sequencing of T-DNA Junctions in Arabidopsis
http://signal.salk.edu/T-DNArecovery.pdf
ECKER LAB
Protocol for nested PCR amplification.

PCR Protocol--In Situ PCR

In Situ PCR

The In Situ PCR:
(Amplification and Detection in a Cellular Context)

Ernest F. Retzel, Katherine A. Staskus, Janet E. Embretson and Ashley T. Haase
Department of Microbiology, University of Minnesota, Minneapolis, MN 55455

Table of Contents
Introduction Technical Aspects/Philosophy Primer Design Reaction Conditions "Mechanics" of the Reaction Methodology
A. Cell and Tissues.
1. Cultured Cells: Infection and Preparation. 2. Tissues: Pulmonary infection and preparation.
B. In Situ Amplification
1. Cultured Cells in Suspension. 2. Tissues on Slides.
C. Detection of Amplified DNA by in situ Hybridization
1. Solution amplification. 2. Tissue Sections.
References

In Situ PCR on Plant Material (Bo Johansen)Required materialsRequired solutionsTissue preparationPectinase treatmentProtease treatmentAcetylationDNase treatmentReverse transcriptionThermal cyclingDetectionMountingAcrobat version of PosterReferencesOther ISPCR linksBotanical Institute

RT IN SITU PCR
Gerard J. Nuovo
TABLE OF CONTENTS
1. Abstract
2. Introductory statement
3. The key preparatory steps
3.1. Fixative
3.1.1. Protease digestion
3.1.2. Definition of optimal protease digestion
3.1.3. Definitions of suboptimal and over-digestion
3.1.4. DNase digestion
3.1.5. Direct incorporation of the reporter nucleotide
4. A one step protocol for RT in situ PCR
5. Applications of RT in situ PCR Matrix metalloprotease expression in cancer as a model for RT in situ PCR
5.1. MMP and TIMP expression in cervical cancer
6. Concluding remarks
7. References
8. Entire manuscript

In situ PCR: protocols and applications.
groups have published protocols and data using in situ PCR techniques. (5). This article will discuss some of the basic concepts of in situ PCR, the proto- ...www.genome.org/cgi/reprint/4/4/S151.pdf
Co-labeling Using In Situ PCR: A Review
Figure 2 Background vs signal with RT in situ PCR. This renal biopsy was analyzed for mRNA by RT in situ PCR. After digestion for 10 min in ...www.jhc.org/cgi/reprint/49/11/1329.pdf
Protocols for the : in situ: PCR-amplification and detection of mRNA and DNA sequencing.
In this protocol we describe the in situ PCR method for the amplification of both DNA and mRNA targets [in situ reverse transcriptase-PCR (RT-PCR)], ...www.nature.com/nprot/journal/v2/n11/abs/nprot.2007.395.html
96-well RNA In Situ Hybridization Protocol
1. Materials. 1.1. Probe Preparation. 1.1.1. Cell Inoculation. 1. 96-well, deep square-well round bottom plate (E&K Scientific Ritter Riplate). ...www.fruitfly.org/about/methods/RNAinsitu.html
A protocol for PCR in situ hybridization of hyphomycetesIntroduction. Within the last years the focus of mycological studies has shifted. emphasis from mere taxonomic studies to more ecological. questions. ...www.im.microbios.org/03setember98/08%20Sterflinger.pdf

PCR Protocol--Tail PCR

TAIL PCR

An introduction of TAIL PCR
Many product bands from the primary TAIL-PCR reaction disappeared after the secondary TAIL-PCR, indicating that these were non-specific type II products ...ihome.cuhk.edu.hk/~z045478/TAIL-PCR%5Bfinal%5D.ppt
TAIL-PCR to clone flanking sequence from Feldmann's linesChun-Ming Liu's HomepageTAIL PCR ANALYSIS
TAIL PCR ANALYSIS. For this service Investigators have three purification options. Our standard purification protocol from the Jackson laboratory, ...www.lajollaneuroscience.org/sr/homepage/dna/tailpcranalysis.html
TAIL PCR ProtocolThe Preuss LabDivision of Biological Sciences, University of Chicago
TAIL PCR
Below is a summary of the responses: I've been using TAIL PCR to isolate the ends of BACs during my chromosome walk. I obtained the sequences of my ...net.bio.net/bionet/mm/arab-gen/1998-September/006602.html
Tail PCR Protocol
Purpose: Mu TAIL PCR produces a population of fragments with one end anchored in Robertson's Mutator (Mu) TIR's and the other in the Mu flanking DNA. ...currant.hos.ufl.edu/mutail/tail.htm
Springer Protocols:High-Throughput TAIL-PCR as a tool to identify DNA flanking insertions.
Springer Protocols is the largest subscription-based electronic database of reproducible laboratory protocols in the Life and Biomedical Sciences.www.springerprotocols.com/Abstract/doi/10.1385/1-59259-413-1:241
TAIL-PCR to clone flanking sequence from Feldmann's lines
The first thing you need to do before starting your TAIL-PCR is to figure out which border of the T-DNA is flanking the plant genome, especially with Ken ...http://www.plant.wageningen-ur.nl/projects/hybtech/Liu/tail_PCR.htm
Isolation of genomic fragment by TAIL-PCR
More than 50bp must be remained downstream of a third gene specific primer to check if a gained TAIL PCR fragment is correctly extended from an original ...www.nibb.ac.jp/evodevo/PHYSCOmanual/4.3.htm

PCR Protocol--Rep PCR

Rep-PCR

The Rep-PCR Genomic Fingerprinting Home PageMichigan State UniversityE. coli Source Tracking by Rep-PCR DNA FingerprintingThe Regents of the University of Minnesota
How does Rep-PCR workThe Rep-PCR Genomic Fingerprinting Home Page
Genomic Fingerprinting by Amplification of Rep-PCR.
Springer Protocols
Rep-PCR DNA Fingerprinting and Computer Analysis (University of Minnesota)
A. Overview (10-25-00)B. Box Fingerprint Protocol (3-4-02)C. Running and Staining Box Fingerprint Gels (10-25-00)
REP-PCR Method Overview (University of Minnesota)
Rep-PCR fingerprinting protocol (University of Minnesota)
https://www.msu.edu/~debruijn/dna5-11.htm

PCR Protocol--Touchdown PCR

Touchdown PCR

Background: Touchdown polymerase chain reaction or touchdown style polymerase chain reaction is a method of polymerase chain reaction by which primers will avoid amplifying nonspecific sequence. The temperature at which primers anneal during a cycle of polymerase chain reaction determines the specificity of annealing. The melting point of the primer sets the upper limit on annealing temperature. At temperatures just below this point, only very specific base pairing between the primer and the template will occur. At lower temperatures, the primers bind less specifically. Nonspecific primer binding obscures polymerase chain reaction results, as the nonspecific sequences to which primers anneal in early steps of amplification will "swamp out" any specific sequences because of the exponential nature of polymerase amplification.
The earliest steps of a touchdown polymerase chain reaction cycle have high annealing temperatures. The annealing temperature is decreased in increments for every subsequent set of cycles (the number of individual cycles and increments of temperature decrease is chosen by the experimenter). The primer will anneal at the highest temperature which is least-permissive of nonspecific binding that it is able to tolerate. Thus, the first sequence amplified is the one between the regions of greatest primer specificity; it is most likely that this is the sequence of interest. These fragments will be further amplified during subsequent rounds at lower temperatures, and will out compete the nonspecific sequences to which the primers may bind at those lower temperatures. If the primer initially binds to the sequence of interest at a low temperature, subsequent rounds of polymerase chain reaction can be performed upon the product to further amplify those fragments.

References
Don R, Cox P, Wainwright B, Baker K, Mattick J (1991). "'Touchdown' PCR to circumvent spurious priming during gene amplification". Nucleic Acids Res 19 (14): 4008. doi:10.1093/nar/19.14.4008. PMID 1861999.
Roux K (1994). "Using mismatched primer-template pairs in touchdown PCR". Biotechniques 16 (5): 812-4. PMID 8068332.
Hecker K, Roux K (1996). "High and low annealing temperatures increase both specificity and yield in touchdown and stepdown PCR". Biotechniques 20 (3): 478-85. PMID 8679209

Berglund:Touchdown PCROpenWetWare
'Touchdown' PCR to circumvent spurious priming during gene amplificationDon RH et al. Nucleic Acids Res. 1991 Jul 25;19(14):4008.
TouchDown PCR
Bio.net
Protocols for PCR and Touchdown PCR:
www.jcsg.org/prod/scripts/primer/pcr_protocol.doc
Combining Multiplex and touchdown PCR for Microsatellite Analysis.
Springer Protocols
Touchdown' PCR to circumvent spurious priming during gene amplification.
Nucleic Acid Res

PCR Protocol--Vectorette PCR

Vectorette PCR

Vectorette End Rescue of Bacterial ClonesStéphanie Le Hellard, University of EdinburghVectorette PCR of mTn-3xHA
The PCR product may then be isolated and sequenced. The vectorette PCR method has the advantage of eliminating transformation and cloning steps. ...ygac.med.yale.edu/mtn/reagent/avail_reagents/vectorette_PCR.html
Vectorette PCR of Yeast DNA
Vectorette PCR of Yeast DNA. Carl Friddle. 1) Cut 1-3 µg of clean DNA overnight with 8-10U of blunt cutting enzyme in 20µl. Most problems come from dirty, ...genomics.princeton.edu/botstein/protocols/vectorette.html
Vectorette PCR Methods
Vectorette PCR was designed for amplification of DNA regions adjacent to known regions ... Performing vectorette PCR with multiple vectorette libraries can ...http://www.bio.psu.edu/People/Faculty/Akashi/vectPCR.html
A Vectorette PCR-based approach for sequencing homologous regions from different genomes.For a successful vectorette PCR, a good specific PCR primer is critical. .... sequencing confirmed that vectorette PCR successfully amplified all expected ...http://www.bio.psu.edu/People/Faculty/Akashi/vect_PCR.online.072304.pdf
Vectorette PCR: a novel approach to genomic walking.
PCR Methods Appl. C Arnold and I J Hodgson. Vectorette PCR: a novel approach to genomic ... genetics method, or vectorette PCR, are. shown in Figure 1. ...www.genome.org/cgi/reprint/1/1/39.pdf
Vectorette PCR isolation of microsatellite repeat sequences using anchored dinucleotide repeat primers
We have developed a vectorette PCR approach to provide an improved method for isolation of microsatellite repeats. The modified procedure relies on PCR ...http://nar.oxfordjournals.org/cgi/content/full/24/11/2190
Springer Protocols:Long Distance Vectorette PCR (LDV PCR).
Springer Protocols is the largest subscription-based electronic database of reproducible laboratory protocols in the Life and Biomedical Sciences.www.springerprotocols.com/Abstract/doi/10.1385/1-59259-177-9:275