TABLE 6.
Alternative primer duplexes to be used for capture of RNAs with specific 3′ ends
| Duplex 3′ overhang | Name | Sequence | RNA strand | Mix ratio | Notes |
|---|---|---|---|---|---|
| +1 G | +G.v3.IR | /5IRD800/GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTG | RNA.v3 or RNA.v3.1 | 1:1 | Excessive levels of adapter-dimer cDNA will accumulate |
| +1 A | +A.v3.IR | /5IRD800/GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTA | RNA.v3 or RNA.v3.1 | 1:1 | |
| +1 T | +T.v3.IR | /5IRD800/GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTT | RNA.v3 or RNA.v3.1 | 1:1 | |
| +1 C | +C.v3.IR | /5IRD800/GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTC | RNA.v3 or RNA.v3.1 | 1:1 | |
| Blunt | +T.v2.IR | /5IRD800/GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT | RNA.v3 or RNA.v3.1 | 1:1 | dG and dA are the most common NTA using current dNTP mix |
| RNA strand | RNA.v3 | rArGrArUrCrGrGrArArGrAmGmCm AmCmAmCmGmUmCmUmGm AmAmCmUmCmCmAmGmU/3SpC3/ | 2′-O-methyl ribonucleotides are immune to RNase H, can potentially interfere with cDNA migration during electrophoresis | ||
| RNA.v3.1 | rArGrArUrCrGrGrArArGrAmGmCmAmCmAmCmGmUrCrUrGrArArCrUmCmCmAmGmUmCmAmC/3SpC3/ | Ribonucleotides introduced between 2′-O-methyl ribonucleotides to ensure sensitivity to RNase H | |||
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Mix ratio represents the molar ratio which the oligonucleotides are combined before annealing. The two listed RNA strand oligonucleotides can be used interchangeably.
