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The binding of proteins to sequences both near to and distant from the promoter can also affect levels of gene expression.

Protein binding can activate transcription by facilitating either RNA polymerase binding or steps farther along in the initiation process, or it can repress transcription by blocking the activity of the polymerase.

In E. coli, one protein that activates transcription is the cAMP receptor protein (CRP), which increases the transcription of genes coding for enzymes that metabolize sugars other than glucose when cells are grown in the absence of glucose. Repressors are proteins that block the synthesis of RNA at specific genes.

In the case of the Lac repressor, transcription of the genes for the enzymes of lactose metabolism is blocked when lactose is unavailable.

greatly increase expression

moderately increase expression

no change in expression

measurably decrease expression

either increase or decrease expression

The rate of expression of the cloned gene is controlled by replacing the gene’s normal promoter and regulatory sequences with more efficient and convenient versions supplied by the vector.

Generally, a well-characterized promoter and its regulatory elements are positioned near several unique restriction sites for cloning, so that genes inserted at the restriction sites will be expressed from the regulated promoter elements.

In some cases, cloned genes are so efficiently expressed that their protein product represents 10% or more of the cellular protein. At these concentrations, some foreign proteins can kill the host cell (usually E. coli ), so expression of the cloned gene must be limited to 24 hours before the planned harvesting of the cells.

Some of these vectors incorporate other features, such as a bacterial ribosome-binding site, to enhance translation of the mRNA derived from the gene or a transcription termination sequence.