S/MARs are constitutive (continuously bound to the matrix or scaffold), or facultative and change their interactions with the matrix depending on cell type or other conditions.

TLS Online TPP Program

#Id: 2640

#Unit 13. Methods in Biology

Run-off transcription is a means of checking the efficiency and accuracy of in vitro transcription.

TLS Online TPP Program

#Id: 2641

#Unit 13. Methods in Biology

The size of this run-off transcript locates the transcription start site, and the amount of this transcript reflects the efficiency of transcription. 

TLS Online TPP Program

#Id: 2642

#Unit 13. Methods in Biology

Notice that S1 mapping and primer extension are well suited to mapping transcripts made in vivo; by contrast, run-off transcription relies on transcription in vitro.

TLS Online TPP Program

#Id: 2643

#Unit 13. Methods in Biology

After you have identified the physiological transcription start site by S1 mapping or primer extension you can use run-off transcription in vitro.

TLS Online TPP Program

#Id: 2644

#Unit 13. Methods in Biology

A variation on the run-off theme of quantifying accurate transcription in vitro is the G-less cassette assay.

TLS Online TPP Program

#Id: 2645

#Unit 13. Methods in Biology

In G-less cassette transcription, a promoter is fused to a double stranded DNA cassette lacking G’s in the nontemplate strand, then the construct is transcribed in vitro in the absence of GTP.