As title suggests, is there a calculated absorbance constant for (ds)DNA:RNA hybrids? i.e. 50 for dsDNA takes the OD260 to ng/uL. Doing this after DNA splint ligation of two RNA fragments to process with DNase and need to know how many ug of RNA there are in solution.
For all practical purposes it will be the same as for ds DNA. However either RNA or DNA in single strand form will be a bit different.
After DNA splint ligation, there is a better way to quantify your ligated RNA product using in situ quantification via gel electrophoresis with the RNA standard (same size of your ligated RNA).
1. Prepare a serial dilution of the RNA standard.
2. Removal of DNA template with DNase I treatment.
3. Load the RNA standard dilutions in the TBE acrylamide gel on a par with your RNA ligation product.
4. Stain the gel with SYBR Gold.
5. Band intensity of your target RNA and the RNA standard can be measured using image analysis software like ImageJ.
Good luck.
@Yun-Tzai Lee Thank you! However, it’s a very expensive synthesis and I’m trying not to waste anything. I even recover the nanodrop volume used.
@Michael J. Benedik Thank you!🙏
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