ARS | Publication request: The role of CPSF30 in 3’ RNA processing of Medicago sativa

Submitted to: Plant and Animal Genome Conference
Publication Type: Abstract Only
Publication Acceptance Date: November 1, 2010
Publication Date: January 15, 2011
Citation: Gaffney, B., Dinkins, R.D., Hunt, A.G. 2011. The role of CPSF30 in 3’ RNA processing of Medicago sativa. Plant and Animal Genome Conference. P412: Legumes, Soybeans, Common Beans.

Technical Abstract: CPSF30 is a component in 3’ RNA processing that has been well characterized in Arabidopsis thaliana. In this study the gene for CPSF30 has been cloned out of Medicago truncatula and Medicago sativa. A large region within the gene exhibits significant conservation between A. thaliana and both Medicago species. This area of the gene was cloned out and 5’ and 3’ RACE were used to get each full length gene. CPSF30 from A. thaliana has been shown to possess three CCCH type zinc fingers, with the first implicated in RNA binding and the third in RNA cleavage. Amino acid alignments showed the zinc finger regions were conserved and it was hypothesized that CPSF30 from both Medicago species would bind RNA and cleave RNA. The M. sativa protein was expressed in E. coli and the purified protein was found to bind RNA. CPSF30 knockouts have been produced in Medicago sativa. Preliminary data suggests that the absence of CPSF30 expression results in a distinct root phenotype. The CPSF30 knockout plants were crossed with a cultivar, Spreader4, which is an agriculturally relevant variety. It has been determined that the knockout vector is present in the hybrid progeny and phenotypes are still being evaluated. High throughput sequencing was used to compare the 3’ ends of the Medicago sativa control and RNAi lines. Preliminary data shows distinct trends in 3’ UTR sequences suggesting that CPSF30 is intricately involved in poly-A track placement.