Species-specific intercistronic primers (rabbit, 5′-AAACAGTAGATGGAGCCTTGATATTC-3′ cow, 5′-CAAATAGTAGATGGAGCCTTGGTG-3′) were then used in 5′ rapid amplification of cDNA ends to recover the full-length SNURF coding sequence. Insert sequences were determined (Macromolecular Resources, Fort Collins, CO). Products were gel-purified and extracted (Qiagen, Chatsworth, CA) and ligated into TA cloning vector pCR2.1 (Invitrogen). These findings identify SNURF as a protein that is produced along with SmN from a bicistronic transcript polycistronic mRNAs therefore are encoded in mammalian genomes where they may form functional operons.Ĭow and rabbit SNURF cDNAs were recovered in two phases by first amplifying the intercistronic region by reverse transcription–PCR of brain poly(A) + mRNA (CLONTECH) with primers corresponding to human and mouse conserved SNURF (RN625: 5′-GGCATTCTTAGCTGAGACACC-3′) and SNRPN (RN626: 5′-ACAATCACAGAGGATCAAATTCAT-3′) coding sequence. We show that both SNURF and SNRPN are translated in normal, but not PWS, human, and mouse tissues and cell lines. Whereas some human tissues express a minor SNURF-only transcript, mouse tissues express only the bicistronic Snurf–Snrpn transcript. Because SNURF is the only protein-coding sequence within the imprinting regulatory region in 15q11–q13, it may have provided the original selection for imprinting in this domain. SNURF encodes a highly basic 71-aa protein that is nuclear-localized (as is SmN).
![macvector chromatogram free macvector chromatogram free](https://image3.slideserve.com/6824475/chromatography3-l.jpg)
Because SNURF–SNRPN maps to human chromosome 15q11–q13 and is paternally expressed, each cistron is a candidate for a role in the imprinted Prader-Willi syndrome (PWS) and PWS mouse models. The vast majority of nucleotide substitutions in SNURF occur in the wobble codon position, providing strong evolutionary evidence for selection for protein-coding function. Phylogenetic analysis of the SNRPN (SmN) mRNA in five eutherian mammals reveals a second highly conserved coding sequence, termed SNURF ( SNRPN upstream reading frame).
![macvector chromatogram free macvector chromatogram free](http://notesforfree.com/wp-content/uploads/2017/09/Chromatography-Sheet-With-Readings.jpg)
Polycistronic transcripts are common in prokaryotes but rare in eukaryotes.