Cite as: Cold Spring Harb. Protoc.; 2008; doi:10.1101/pdb.prot4858

This Protocol
Right arrow Full Text
Right arrow Update/discuss this protocolDiscussion icon
Right arrow Alert me when this protocol is cited
Right arrow Alert me when comments are published
Right arrow Alert me if a correction is posted
Services
Right arrow Similar protocols in this database
Right arrow Alert me to new releases of protocols
Right arrow Save to Personal Folders
Right arrow Download to citation manager
Right arrow Printer-friendly versionPrinter-friendly version
Citing Articles
Right arrow Citing Articles via HighWire
Google Scholar
Right arrow Articles by Rebrikov, D. V.
PubMed
Right arrow Articles by Rebrikov, D. V.
Related Collections
Right arrow Cell Biology, general
Right arrow Molecular Biology, general
Right arrow Analysis of Gene Expression
Right arrow Analysis of Gene Expression, general
Right arrow cDNA
Right arrow Libraries
Right arrow Libraries, general
Right arrow cDNA Libraries
Right arrow Genomic Libraries
Right arrow RNA
Right arrow RNA, general
Right arrow Analysis of Gene Expression in Cultured Cells
Right arrow Expression Libraries
Right arrow Polymerase Chain Reaction (PCR)
Right arrow Polymerase Chain Reaction (PCR), general
Right arrow Differential Gene Expression
Right arrowRelated Article
Right arrowRelated Protocols
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?
Legend icon

protocolProtocol

Identification of Differential Genes by Suppression Subtractive Hybridization: IV. Mirror Orientation Selection (MOS)

Denis V. Rebrikov

This protocol was adapted from "Identification of Differential Genes by Suppression Subtractive Hybridization," Chapter 22, in PCR Primer, 2nd edition, (eds. Dieffenbach and Dveksler). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2003.


INTRODUCTION

Suppression subtractive hybridization (SSH) is one of the most powerful and popular methods for generating subtracted cDNA or genomic DNA libraries. This technique can be used to compare two mRNA populations and obtain cDNAs representing genes that are either overexpressed or exclusively expressed in one population as compared to another. It can also be used for comparison of genomic DNA populations. The major drawback of SSH is the presence of background clones that represent nondifferentially expressed DNA species in the subtracted libraries. In some cases, the number of background clones may considerably exceed the number of target clones. This protocol describes mirror orientation selection (MOS)--a simple procedure that substantially decreases the number of background clones in libraries generated by SSH. The MOS technique is based on the rationale that after PCR amplification, during SSH, background molecules will be present in one orientation only, relative to the adapter sequences. Genuine SSH clones will be present in both sequence orientations.


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?

Related Article

Identification of Differential Genes by Suppression Subtractive Hybridization: An Overview
Denis V. Rebrikov
CSH Protocols 2008: 21. [Abstract] [Full Text]

Related Protocols

Purification of Nucleic Acids by Extraction with Phenol:Chloroform
Joseph Sambrook and David W. Russell
CSH Protocols 2006: 4455. [Extract] [Full Text]

Standard Ethanol Precipitation of DNA in Microcentrifuge Tubes
Joseph Sambrook and David W. Russell
CSH Protocols 2006: 4456. [Extract] [Full Text]

Agarose Gel Electrophoresis
Joseph Sambrook and David W. Russell
CSH Protocols 2006: 4020. [Extract] [Full Text]

Identification of Differential Genes by Suppression Subtractive Hybridization: I. Preparation of Subtracted cDNA or Genomic DNA Library
Denis V. Rebrikov
CSH Protocols 2008: 4855. [Abstract] [Full Text]

Identification of Differential Genes by Suppression Subtractive Hybridization: II. Subtractive Hybridization
Denis V. Rebrikov
CSH Protocols 2008: 4856. [Abstract] [Full Text]

Identification of Differential Genes by Suppression Subtractive Hybridization: III. PCR Amplification of Differentially Presented DNAs
Denis V. Rebrikov
CSH Protocols 2008: 4857. [Abstract] [Full Text]

Identification of Differential Genes by Suppression Subtractive Hybridization: V. PCR-Based DNA Dot Blot
Denis V. Rebrikov
CSH Protocols 2008: 4859. [Abstract] [Full Text]

Identification of Differential Genes by Suppression Subtractive Hybridization: VI. Differential Hybridization with Tester and Driver DNA Probes
Denis V. Rebrikov
CSH Protocols 2008: 4860. [Abstract] [Full Text]



This article has been cited by other articles:


Home page
 CSH ProtocolsHome page
D. V. Rebrikov
Identification of Differential Genes by Suppression Subtractive Hybridization: An Overview
CSH Protocols, July 1, 2008; 2008(8): pdb.top21 - pdb.top21.
[Abstract] [Full Text]

Home page
 CSH ProtocolsHome page
D. V. Rebrikov
Identification of Differential Genes by Suppression Subtractive Hybridization: I. Preparation of Subtracted cDNA or Genomic DNA Library
CSH Protocols, July 1, 2008; 2008(8): pdb.prot4855 - pdb.prot4855.
[Abstract] [Full Text]

Home page
 CSH ProtocolsHome page
D. V. Rebrikov
Identification of Differential Genes by Suppression Subtractive Hybridization: II. Subtractive Hybridization
CSH Protocols, July 1, 2008; 2008(8): pdb.prot4856 - pdb.prot4856.
[Abstract] [Full Text]

Home page
 CSH ProtocolsHome page
D. V. Rebrikov
Identification of Differential Genes by Suppression Subtractive Hybridization: III. PCR Amplification of Differentially Presented DNAs
CSH Protocols, July 1, 2008; 2008(8): pdb.prot4857 - pdb.prot4857.
[Abstract] [Full Text]

Home page
 CSH ProtocolsHome page
D. V. Rebrikov
Identification of Differential Genes by Suppression Subtractive Hybridization: V. PCR-Based DNA Dot Blot
CSH Protocols, July 1, 2008; 2008(8): pdb.prot4859 - pdb.prot4859.
[Abstract] [Full Text]

Home page
 CSH ProtocolsHome page
D. V. Rebrikov
Identification of Differential Genes by Suppression Subtractive Hybridization: VI. Differential Hybridization with Tester and Driver DNA Probes
CSH Protocols, July 1, 2008; 2008(8): pdb.prot4860 - pdb.prot4860.
[Abstract] [Full Text]