In Table 1, there are missing column headers. Please see the complete, correct Table 1 here.
Table 1. RP-HPLC quantification of nucleosides in S.cerevisiae 18S and 25S rRNA.
| 18S rRNA | 25S rRNA | |||
|---|---|---|---|---|
| Nucleosides | Mole % (Seq) | Mole % (HPLC) | Mole % (Seq) | Mole %(HPLC) |
| C (Cytidine) | 19.1 | 19.5 ± 0.48 | 19.2 | 19.4 ± 0.56 |
| U (Uridine) | 27.4 | 27.1 ± 0.30 | 24.4 | 24.9 ± 0.39 |
| G (Guanosine) | 25.2 | 25.7 ± 0.12 | 28.2 | 27.5 ± 0.17 |
| A (Adenosine) | 26.3 | 26.6 ± 0.68 | 26.1 | 26.8 ± 0.64 |
| Residues/mole |
Residues/mole (HPLC) |
Residues/mole |
Residues/mole (HPLC) |
|
| m1A (N1-Methyladenosine) | 0 | 0 | 2 | 1.98 ± 0.10 |
| m5C (C5-Methylcytidine) | 0 | 0 | 2 | 1.81 ± 0.17 |
| Cm (2’-O-Methylcytidine) | 3 | 3.1 ± 0.26 | 7 | 6.74 ± 0.31 |
| m7G (N7-Methylguanosine) | 1 | 0.72*± 0.16 | 0 | 0 |
| m5U (C5-Methyluridine) | 0 | 0 | 0 | 0 |
| Um (2’-O-Methyluridine) | 2 | 1.89 ± 0.33 | 8 | 7.1 ± 0.31 |
| m3U (N3-Methyluridine) | 0 | 0 | 2 | 2.1 ± 0.17 |
| Gm (2’-O-Methylguanosine) | 5 | 4.91 ± 0.27 | 10 | 9.3 ± 0.39 |
| ac4C (N4-Acetylcytidine) | 2 | 2.12 ± 0.11 | 0 | 0 |
| Am (2’-O-Methyladenosine) | 8 | 7.33 ± 0.43 | 12 | 11.8 ± 0.48 |
| m62A (N6,N6-Dimethyladenosine) | 2 | nc | 0 | 0 |
Mole % (Seq) was calculated from the number of each nucleoside in 18S and 25S rRNA sequences and as explained in Materials and Methods. Mole % (HPLC), Residues/mole (HPLC) and standard deviation values were calculated from the peak area of the respective nucleosides using their standard response factors, from nine independent runs for each 18S and 25S rRNA.
nc—not calculated
* m7G has been shown to undergo partial-degradation during hydrolysis [18].
There are several errors in Table 2. Please see the correct Table 2 here.
Table 2. Approximate values for the extent of modifications in 18S and 25S rRNA, using RP-HPLC and its comparison with RiboMethSeq and SILNAS analyses.
| Modified residue | Made by | % of modification (RP-HPLC) |
% of modification (Ribo-Meth Seq) (Ref.35) |
% of modification (Ribo-Meth Seq) (Ref.34) |
% of modification (SILNAS) (Ref.37) |
|---|---|---|---|---|---|
| 18S rRNA | |||||
| Am28 | snR74 | 90% ± 3.8% | 85% | 99% | >95% |
| Am100 | snR51 | 73% ± 2.4% | 77% | 77% | 80% |
| Cm414 | U14 | 89% ± 3.1% | 89% | 96% | >95% |
| Am420 | snR52 | 98% ± 1.5% | 83% | 90% | >95% |
| Am436 | snR87 | 98% ± 1.8% | 74% | 89% | 73% |
| Am541 | snR41 | 95% ± 3.2% | 87% | 99% | >95% |
| Gm562 | snrR40 | 100% | 94% | 92% | 67% |
| Um578 | snR77 | 95% ± 3.3% | 90% | 93% | >95% |
| Am619 | snR47 | 98% ± 1.2% | 85% | 92% | 100% |
| Am796 | snR53 | 98% ± 1.1% | 92% | 97% | >95% |
| Am974 | snR54 | 95% ± 3.4% | 91% | 96% | 94% |
| Cm1007 | snR79 | 100% | 92% | 98% | >95% |
| Gm1126 | snR41 | 100% | 94% | 93% | 89% |
| m1acp3Ψ1191 | snR35-Nep1-Tsr3 | nd | nd | nd | 100% |
| Um1269 | snR55 | 95% ± 2.1% | 88% | 92% | >95% |
| Gm1271 | snR40 | 100% | 92% | 98% | >95% |
| ac4C1280 | Kre33 | 96% ± 1.9% | nd | nd | 84% |
| Gm1428 | snR56 | 100% | 95% | 99% | >95% |
| Gm1572 | snR57 | 100% | 89% | 97% | 100% |
| m7G1575 | Bud23 | 72*% ± 2.8% | nd | nd | >95% |
| Cm1639 | snR70 | 92% ± 2.9% | 93% | 98% | 79% |
| ac4C1773 | Kre33 | 100% | nd | nd | >95% |
| m62A1781 | Dim1 | nc | nd | nd | 90% |
| m62A1782 | Dim1 | nc | nd | nd | 90% |
| 25S rRNA | |||||
| m1A645 | Rrp8 | 99% ± 1% | nd | nd | >95% |
| Am649 | U18 | 93% ± 2.5% | 88% | 98% | >95% |
| Cm650 | U18 | 100% | 95% | 98% | 94% |
| Cm663 | snR58 | 81% ± 4.3% | 62% | 91% | 74% |
| Gm805 | snR39b | 90% ± 4.7% | 93% | 98% | >95% |
| Am807 | snR39_snR59 | 98% ± 1% | 94% | 97% | >95% |
| Am817 | snR60 | 98% ± 0.8% | 92% | 85% | 89% |
| Gm867 | snR50 | 90% ± 3.7% | 95% | 95% | 78% |
| Am876 | snR72 | 95% ± 2.2% | 89% | 91% | 75% |
| Um898 | snR40 | 94% ± 1.8% | 78% | 85% | 94% |
| Gm908 | snR60 | 95% ± 1.1% | 96% | 96% | 100% |
| Am1133 | snR61 | 98% ± 0.5% | 92% | 99% | 88% |
| Gm1142 | ?(snr57) | absent | absent | absent | absent |
| Cm1437 | U24 | 100% | 92% | 97% | >95% |
| Am1449 | U24 | 97% ± 0.7% | 67% | 99% | >95% |
| Gm1450 | U24 | 100% | 94% | 99% | >95% |
| Um1888 | snR62 | 99% ± 1.3% | 94% | 97% | >95% |
| m1A2142 | Bmt2 | 99% ± 0.7% | nd | nd | >95% |
| Cm2197 | snR76 | 100% | 71% | 86% | 91% |
| Am2220 | snR47 | 98% ± 3.1% | 94% | 99% | 93% |
| Am2256 | snR63 | 98% ± 2.3% | 94% | 95% | 100% |
| m5C2278 | Rcm1 | 100% | nd | nd | 100% |
| Am2280 | snR13 | 99% ± 0.6% | 91% | 100% | 100% |
| Am2281 | snR13 | 99% ± 0.9% | 92% | 100% | 100% |
| Gm2288 | snR75 | 100% | 95% | 96% | >95% |
| Cm2337 | snR64 | 100% | 94% | 99% | 100% |
| Um/Ψm 2347 | snR65/snR9 | 66% ± 3.7% | 69% | 61% | 76.2%/12.9% |
| Gm2395 | ?snR190 | absent | absent | absent | absent |
| Um2417 | snR66 | 99% ± 0.1% | 95% | 99% | 100% |
| Um2421 | snR78 | 99% ± 1.5% | 86% | 93% | >95% |
| Gm2619 | snR67 | 100% | 93% | 98% | 87% |
| m3U2634 | Bmt5 | 100% | nd | nd | >95% |
| Am2640 | snR68 | 95% ± 3.6% | 90% | 96% | 93% |
| Um2724 | snR67 | 92% ± 2.1% | 92% | 98% | >95% |
| Um2729 | snR51 | 89% ± 4.3% | 67% | 94% | 75% |
| Gm2791 | snR48 | 95% ± 2.3% | 94% | 98% | 86% |
| Gm2793 | snR48 | 95% ± 1.9% | 95% | 98% | >95% |
| Gm2815 | snR38 | 95% ± 2% | 95% | 99% | >95% |
| m3U2843 | Bmt6 | 100% | nd | nd | >95% |
| m5C2870 | Nop2 | 100% | nd | nd | >95% |
| Um2921 | snR52 | 93% ± 2.2% | 92% | 99% | >95% |
| Gm2922 | Spb1 | 97% ± 3.5% | 95% | 98% | >95% |
| Am2946 | snR71 | 99% ± 0.6% | 92% | 99% | 91% |
| Cm2948 | snR69 | 99% ± 0.8% | 77% | 82% | >95 |
| Cm2959 | snR73 | 93% ± 4.1% | 96% | 95% | 94% |
nd—not detected; nc—not calculated.? (snRx)- predicted according to the guide sequence; lacks experimental evidence
% modification was calculated as a mean of three independent calculations ± Relative standard deviations (RSD).
m7G has been shown to undergo partial-degradation during hydrolysis [18].
Reference
- 1.Yang J, Sharma S, Watzinger P, Hartmann JD, Kötter P, Entian K-D (2016) Mapping of Complete Set of Ribose and Base Modifications of Yeast rRNA by RP-HPLC and Mung Bean Nuclease Assay. PLoS ONE 11(12): e0168873 doi: 10.1371/journal.pone.0168873 [DOI] [PMC free article] [PubMed] [Google Scholar]