DNA footprint protection (described in Research Technique 8.1) is a method that determines whether proteins bind to a specific sample of DNA and thus protect part of the DNA from random enzymatic cleavage by DNase I. A 400-bp segment of cloned DNA is thought to contain a promoter. The cloned DNA is analyzed by DNA footprinting to help determine if it has the capacity to act as a promoter sequence. The accompanying gel has two lanes, each containing the cloned 400-bp DNA fragment treated with DNase I to randomly cleave unprotected DNA. Lane 1 is cloned DNA that was mixed with RNA polymerase II and several TFII transcription factors before exposure to DNase I. Lane 2 contains cloned DNA that was exposed only to DNase I. RNA pol II and TFIIs were not mixed with that DNA before adding DNase I. Explain why this gel provides evidence that the cloned DNA may act as a promoter sequence.

Key Concepts

DNA Footprinting

DNA footprinting is a technique used to identify the specific regions of DNA that are bound by proteins, such as transcription factors. In this method, DNA is treated with an enzyme like DNase I, which cleaves unprotected DNA. The resulting fragments are analyzed through gel electrophoresis, allowing researchers to visualize which parts of the DNA remain intact, indicating protein binding sites.

Promoter Sequences

Promoter sequences are specific regions of DNA that initiate transcription of a gene. They are typically located upstream of the coding region and are recognized by RNA polymerase and transcription factors. The presence of a promoter is crucial for the binding of RNA polymerase, which is necessary for the synthesis of RNA from the DNA template.

Transcription Factors

Transcription factors are proteins that bind to specific DNA sequences to regulate the transcription of genes. They can enhance or inhibit the recruitment of RNA polymerase to the promoter region. In the context of the gel analysis, the presence of RNA polymerase II and transcription factors in Lane 1 suggests that they are binding to the cloned DNA, protecting it from DNase I cleavage and indicating the potential of that DNA to function as a promoter.