Part:BBa_K2429012

pENTR Ms2

This part produces a RNA bacteriophage Ms2 coat protein. This protein is responsible for extending the life of the virus by binding to stem loops on the RNA, and allows the genetic information to be encapsidated in the protein [1]. A stem loop is a secondary structure of RNA that occurs when the RNA folds in on itself, and two complimentary bases form a bond and results in a U-shaped loop forming along the strand. The Ms2 protein is also involved in reducing the translation of replicase protein. The replicase protein uses the genetic RNA as a template to make double stranded RNA. The bacteriophages displaying Ms2 are capable of reproducing its genetic RNA independent of primers; however, it still requires the RNA as single-stranded templates. With excess replicase, the RNA would be in double-stranded form, and the virus unable to replicate the RNA [2]. Hence, strengthening the importance of Ms2 coat protein to virus survival.

Our team used this part to attempt to control alternative splicing. Typically, during the process of alternative splicing in eukaryotic cells, specific RNA binding proteins will bind to sequences or motifs on a pre-mRNA strand and eventually come together to form a spliceosome protein. This resulting protein will cleave the mRNA strand to exclude portions of the pre-mRNA (called introns) and the remaining sequences in the mature mRNA are known as exons. Our team used this protein in an attempt to control what exons would be included in an mRNA transcript by targeting the motifs in an intron, thus blocking splicing factors from binding and retaining an exon

[1]Peabody, D S. “The RNA Binding Site of Bacteriophage MS2 Coat Protein.” The EMBO Journal 12.2 (1993): 595–600. Print.

[2]http://what-when-how.com/molecular-biology/qb-replicase-molecular-biology/

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 294
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 294
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 294
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 294
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 294
  
• 1000
  COMPATIBLE WITH RFC[1000]