PCR Primer Design

Definition PCR primer design is the creation of short nucleotide sequences for use in amplifying a specific region of DNA.

Examples PCR primers are designed to:  Highly conserved DNA regions  Protein-coding regions with low degeneracy  More conserved regions that flank variable regions

Non-examples PCR primers are not designed to:  Repeat regions  Regions with secondary structure  Regions that can form primer-dimers  Regions with low G/C content

Applications Primer design is used for:  Finding new genes  Developing new identification tools  Optimizing PCRs

Primer Design Criteria Target: Conserved nucleotide or protein regions Length: Usually bases Purine:pyrimidine content of around 1:1 End with 1-2 GC pairs, if possible No inter- or intra-primer interactions Check with databases for specificity Cycling conditions and buffer concentrations should be adjusted for each primer pair (see PCR troubleshooting)

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California State Chemistry Standards Grade 8 6c. Living organisms have many different kinds of molecules… Grades Solutions are homogenous mixtures of two or more substances 7. Energy is exchanged or transformed in all chemical reactions and physical changes of matter 8. Chemical reaction rates depend on factors that influence the frequency of collision of reactant molecules

California State Biology Standards Grade 7 2e. DNA is the genetic material of living organisms and is located in the chromosomes of each cell 3a. Genetic variation and environmental factors are causes of evolution and diversity of organisms Grades Mutation and sexual reproduction lead to genetic variation in a population 7. The frequency of an allele in a gene pool of a population depends on many factors and may be stable or unstable over time

California State Investigation and Experimentation Standards Grades 7, 9-12 a/b. Select and use appropriate tools and technology to perform tests, collect data, analyze relationships, and display data Grade 8 a. Plan and conduct a scientific investigation to test a hypothesis

National Standards Grades 6-12 Content Standard A: Science as Inquiry Content Standard C: Life Science Content Standard E: Science and Technology

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