RNA

Part:BBa_K3602002:Design

Designed by: Flora Fuglsang   Group: iGEM20_SDU-Denmark   (2020-10-23)


ERG sgRNA Cas13a


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

For the design of this sgRNA, the targeted gene was first identified from the human reference genome GRCh38 via ensembl (https://www.ensembl.org/). The specific sequence was chosen as 28 nucleotides which were present in an exon of most of the possible splicing variants. The found sequence is used directly in the sgRNA being the spacer. After in-vitro transcription, the sequence will be the reverse complementary and thus able to bind to the targeted biomarker.

The scaffold sequence was designed to match the requirements for cas13a according to Gootenberg et al. 2017. (https://science.sciencemag.org/content/356/6336/438)

The T7 promoter was chosen from neb.com for high yield RNA synthesis. (https://international.neb.com/protocols/2015/11/24/sgrna-synthesis-using-the-hiscribe-quick-t7-high-yield-rna-synthesis-kit-neb-e2050)

Source

The sequence is designed from the ERG mRNA sequence from the human reference genome GRCh38 via ensembl (https://www.ensembl.org/).

References