Measurement

Part:BBa_K3014009:Design

Designed by: Katharina Hofer, Sini Münßinger, Jan Seeger, Gabriele Kepp   Group: iGEM19_Stuttgart   (2019-10-20)


DNA-RNA-Hybrid-Adapter for tRNA quantification


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 121
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 121
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 121
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 121
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 121
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Therefore, we designed a linear DNA/RNA hybrid adapter, that specifically binds to the 5’ terminus of mature tRNA.It is complementary to the 3’ overhang of tRNA after the amino acid was removed by diacylation treatment. Using a forward primer, that binds to the adapter and a tRNA specific reverse primer, the desired tRNA can be amplified in a PCR reaction. Individual tRNA levels can be quantified with the use of a DNA intercalating dye.

The adapter is composed of DNA except of the last two 3’-terminal nucleotides, which are composed of RNA. All tRNA molecules show an identical sequence at their 3’ terminus: ACCN, to which the adapter anneals. The last 3’-terminal nucleotide of the adapter should be complementary to the descriminator base of the respective tRNA.


Source

This part was synthesized by Integrated DNA Technologies.