Previous studies in vivo reported that processing of primary microRNA 
(pri-miRNA) is coupled to transcription by RNA polymerase II (RNAP II) and can 
occur co-transcriptionally. Here we have established a robust in vivo system in 
which pri-miRNA is transcribed by RNAP II and processed to pre-miRNA in HeLa 
cell nuclear extracts. We show that both the kinetics and efficiency of 
pri-miRNA processing are dramatically enhanced in this system compared to that 
of the corresponding naked pri-miRNA. Moreover, this enhancement is general as 
it occurs with multiple pri-miRNAs. We also show that nascent pri-miRNA is 
efficiently processed before it is released from the DNA template. Together, our 
work directly demonstrates that transcription and pri-miRNA processing are 
functionally coupled and establishes the first in vivo model systems for this 
functional coupling and for co-transcriptional processing.