I cloned my designed gene fragment in L4440 vector under the control of two T7-promoters to get a double stradouble-strandednded RNA (dsRNA). This cloned vector is transformed into an HT115 RNase-deficient E. coli. To isolate dsRNA, then I extracted the total nucleic acid of HT115 RNase-deficient E. coli. Then nucleic acid was treated with RNase A (Thermo Scientific) to remove single single-stranded RNA in presence of 0.5 M NaCl and 10 ng of Rnase A for 1000 ng of RNA, incubated for 30 minutes at 37°C. observed visualized RNase A treated samples in agarose gel I could not observe anything. All degraded. RNase A treatment also digests dsRNA.
Kindly help to me to purify intact dsRNA from HT115 RNase-deficient E. coli.
refer to previous discussion
https://www.researchgate.net/post/How_to_purify_dsRNA_produced_by_E_coli2
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