1 / 13

DNA Microarray:

DNA Microarray:. A Recombinant DNA Method. Basic Steps to Microarray:. Obtain cells with genes that are needed for analysis. Isolate the mRNA using extraction buffer. Remove the buffer with RNA. The mRNA will be 5’-3’ with repeating AAA..end. Convert each mRNA into colored cDNA.

melina
Download Presentation

DNA Microarray:

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. DNA Microarray: A Recombinant DNA Method

  2. Basic Steps to Microarray: • Obtain cells with genes that are needed for analysis. • Isolate the mRNA using extraction buffer. • Remove the buffer with RNA. • The mRNA will be 5’-3’ with repeating AAA..end. • Convert each mRNA into colored cDNA. • Targets labeled with fluorophores.

  3. Basic Steps to Microarray: • With mRNA degraded reverse cDNA created. • Mix colored samples of cDNA.

  4. Microarray or DNA Chip: • Every spot on the chip represents a different coding sequence from different genes. • Each spot on the chip is made of a DNA probe that can pair with the cDNA that was created.

  5. Basic Steps to Microarray: • Incubation with the mixed cDNA and the chip DNA will yield some pairing. • Wash off unbound cDNA to see what has bound to the microarray.

  6. Visualizing Bound cDNA: • The slide with the microarray chip is placed inside a dark box where it is scanned with a high resolution laser that detects the bound fluorescent labels. • The information and images are then sent to the computer for analysis.

  7. Analyzing the Data: • Creates a ratio image. • Green images signal expression in one condition. • Red images signal expression in one condition. • Yellow images signal expression in both conditions.

  8. Capacities: • Not limited to human genetic material. • Can be used for all species. • Can display thousands of different genes. • Pending chip size. • 100’s nucleotides (<1 cm^2) • 100,000’s nucleotides (about 10 cm^2) • Allows the study of multiple genes at once.

  9. Types of Microarray: • Pre-synthesized nucleic acids. • Bought cDNA. • Oligonucleotide synthesis in situ. • Creating cDNA in question.

  10. Microarray Application: • Identifying gene’s and gene mutations of different types of diseases such as cancer. • Identifying the expression level, or quantity of genes (mutated or not).

  11. Definitions: • Target - the nucleic acid (cDNA) sample who’s identity and quantity are being measured. • Fluorophore – usually green and red labels attached to the target to enable visualizing expression. • Microarray works as reverse hybridization method converting from mRNA to cDNA 3’-5’ with TTTT…end. • Probe – an attached nucleic acid with a known sequence (the DNA chip).

  12. Resources: • Strachan, Tom; Read, Andrew. Human Molecular Genetics 3. pp 175-178. New York. Garland Publishing. 2004. • Campbell,A. Malcom. Microarray Animation. Davidson College. 2001. www.bio.davidson.edu/courses/genomics/chip/chip.html • http://cmgm.stanford.edu • http://www.gene-chips.com/

More Related