JP2750127B2 - Method for Cleavage of Specific Site of Polyribonucleotide by Ribozyme - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to a polyribonucleotide constituting a ribozyme having a catalytic activity for cleaving a specific site in a polyribonucleotide sequence, and also relates to the catalytic polyribonucleotide. The present invention relates to a method for cleaving a specific site in another polyribonucleotide sequence using nucleotides.

As an example of industrial use of the present invention, there is expected a possibility of cutting and detoxifying viroid single-stranded cyclic RNA, which is considered to be a cause of dwarfing of various plants.

(Prior Art) There have been several reports on RNA or polyribonucleotides having self-cleaving activity. Zaug et al.
We found that the precursor liposomal RNA of Tetrahymena forms ribosomal RNA by self-cleavage (Nature, 324 ,
429-433, (1986)). Hutchins and colleagues found that the positive and negative RNA strands of avocado sunbloch viroid each self-cleave, and that the nucleotide sequence near the cleavage site was highly homologous,
The RNA strand has a similar secondary structure near the cleavage site, and it was predicted that the cleavage would occur at a specific site in the specific sequence (Nu
cleic Acid Res. 14, 3627-3640, (1986)).

Uhlenbeck uses T7 RNA polymerase and synthetic DNA,
24mer and 19mer containing the consensus sequence of this viroid
And found that self-cleavage occurs at a specific site of the 24mer (Nature, 328 , 5
96-600, (1987). Epstain et al. Found that a transcript with a chain length of about 300 of newt satellite DNA self-cleaves at a specific site, and the nucleotide sequence near the cleavage site was similar to the nucleotide sequence near the cleavage site of Avogado Sanbrotch viroid. (Cell, 48 , 535-543, (1987)).

First, the present inventors chemically synthesized two types of 21-mer polyribonucleotides containing a cleavage site of this newt satellite RNA, and when a double strand was formed for this, one strand was cleaved at a specific site. (FEBS L
ett., 228 , 228-230, (1988)). At the same time, sequences essential for self-cleavage were examined, and 9-14 AU and 10-13 C
-G base pairs were found to be interchangeable. These RNAs, RNAs having polyribonucleotide cleavage activity, and polyribonucleotides are collectively referred to as ribozymes.

However, all ribozymes found hitherto are capable of self-cleaving, and none have been found to cleave other RNA or polyribonucleotide molecules.

(Problems to be Solved by the Invention) The present inventors have studied a polyribonucleotide sequence that recognizes a specific sequence of another polyribonucleotide without performing self-cleavage and cleaves a specific site. As a result, two-dimensionally different polyribonucleotides of two different sequences, or polyribonucleotides having the two types of sequences in the same molecule, recognizing a polyribonucleotide having a specific sequence in a three-dimensional structure Finding a method for cleaving a polyribonucleotide having the specific sequence,
The present invention has been completed. That is, the present invention relates to a polyribonucleotide including a specific sequence constituting a ribozyme having a function of cleaving another polyribonucleotide at a specific site, and further relates to the polyribonucleotide and another specific sequence. The present invention relates to a method for cleaving a specific site of a third polyribonucleotide having a specific sequence by constituting a ribozyme with the polyribonucleotide having the ribozyme.

The present invention relates to a polyribonucleotide comprising the nucleotide sequence represented by A of the formula (b) and not comprising the nucleotide sequence represented by C;
Using a polyribonucleotide not containing the nucleotide sequence represented by A, or using a polyribonucleotide containing the nucleotide sequence represented by A and the nucleotide sequence represented by B and not containing the nucleotide sequence represented by C , C
The present invention relates to a method for cleaving a polyribonucleotide containing a nucleotide sequence represented by the following formula: : (In the formula, U represents a uracil nucleotide, A represents an adenine nucleotide, C represents a cytosine nucleotide, G represents a guanine nucleotide, R represents any of a uracil nucleotide, an adenine nucleotide, and a cytosine nucleotide, and corresponds to S _{3 to} S ₅ . Y ₁ corresponding to X _{3 to} X ₅ , V _{1 to} V ₂
~ Y ₂ , and Z _{3 to} Z ₅ corresponding to W _{3 to} W ₅ are uracil nucleotides, adenine nucleotides, which can form complementary hydrogen bonds, respectively, of which the integer numbers in the combination of these three groups match. cytosine nucleotides, represent either guanine nucleotide, W ₁ corresponding to _{X 1 ~X 2, V 3 ~V} 4 and the corresponding Y ₃ to Y _4, and Z ₁ to Z ₂ and the corresponding S ₁ to S ₂ ~ W ₂ is a uracil nucleotide, an adenine nucleotide, a cytosine nucleotide, in which at least two of the combinations of the three groups can form complementary hydrogen bonds, each having the same integer number,
Represents any of the guanine nucleotides. ).

In particular, in formula (b), X ₁ ~X ₂ and the corresponding S ₁ to S _2,
V ₃ ~V ₄ and the corresponding Y ₃ to Y _4, and Z ₁ to Z ₂ corresponds to the W ₁ to _W-2
The present invention relates to a method for cleaving a polyribonucleotide, which is any one of uracil nucleotides, adenine nucleotides, cytosine nucleotides, and guanine nucleotides, each of which has an identical integer number in a combination of these three groups and can form a complementary hydrogen bond.

Particularly preferably, in the formula (b), R, S ₁ , W ₁ ,
W ₃ , X ₅ , Y ₃ , Z ₂ and Z ₅ are adenine nucleotides, S ₂ ,
S ₃ , S ₄ , V ₂ , W ₄ , Y ₁ and Y ₄ are cytosine nucleotides,
V ₁ , V ₄ , X ₂ , X ₃ , X ₄ , Y ₂ and Z ₄ are guanine nucleotides, and S ₆ , V ₃ , W ₂ , W ₅ , X ₁ , Z ₁ and Z ₃ are uracil nucleotides The present invention relates to a method for cleaving a polyribonucleotide.

Then, the A-chain of the formula (b) is a polyribonucleotide having a nucleotide sequence represented by the formula (I): (Where U represents a uracil nucleotide, A represents an adenine nucleotide, C represents a cytosine nucleotide, and G represents a guanine nucleotide), which is a novel polyribonucleotide, and is particularly preferred.

(Means for Solving the Problems) A) Synthesis of Polyribonucleotide Chain Polyribonucleotide chain is synthesized by condensing unit nucleotides by solid phase phosphoramidite method after protecting 2'-hydroxyl group and 5'-hydroxyl group. (Seismic Chemistry Experiment Course 1, Genetic Research Method I, pp.35-6)
1, 1986, Tokyo Doujinshi). That is, a phosphoramidite is reacted with a nucleoside in which a 2'-hydroxyl group is protected by a tetrahydrapyranyl group or the like, a 5'-hydroxyl group is protected by a dimethoxytrityl group or the like, and the base is protected as necessary. Thus, a completely protected unit nucleotide (nucleoside 3'-0-phosphoramidite) can be obtained. The extension of the polyribonucleotide chain is caused by the 5'-hydroxyl group of the nucleoside resin containing the target base in which the 5'-hydroxyl group is protected by a dimethoxytrityl group or the like and the 3'-hydroxyl group is bonded to the carrier via a crosslinked structure. It can be carried out by removing the protecting group and sequentially reacting the desired unit nucleotide.

B) Removal and Purification of Protecting Group Next, removal of the methyl group attached to the phosphate group, cleavage of the polyribonucleotide chain from the resin by alkali treatment, removal of the protecting group at the base moiety, and 2′-acid treatment by acid treatment
The target polyribonucleotide chain can be obtained by removing the protecting group for the hydroxyl group and the protecting group for the 5′-hydroxyl group, followed by a desalting treatment, a purification operation using reverse phase, ion exchange chromatography and the like.

C) Confirmation of the base sequence of the polyribonucleotide chain To confirm the base sequence of the polyribonucleotide chain, label the 5 'end of the polyribonucleotide chain, fragment it with a snake venom phosphodiesterase, for example, electrophoresis and homochromatography. This can be done by performing lithography, etc.
-29, 1986, Tokyo Chemical Dojin). The chain length can also be determined by performing electrophoresis on the labeled polyribonucleotide chain.

D) Cleavage of other polyribonucleotide chains Cleavage of other polyribonucleotide chains by polyribonucleotide chains can be confirmed by the following operations (chemical,
Vol. 43, No. 5, pp. 348-349 (1988)). The 5'-end of the polyribonucleotide chain used as a substrate is labeled, and a buffer containing a polyribonucleotide chain having a catalytic activity, magnesium chloride, and sodium chloride is added thereto, and the mixture is kept warm. After a certain time, the reaction is stopped by adding EDTA to the reaction solution, and the solution is subjected to polyacrylamide gel electrophoresis. After the electrophoresis, the gel is cut out, and the radioactivity at the cleavage site is quantified to calculate the cleavage rate.

Hereinafter, as a reference example, a method for synthesizing a nucleoside 3'-0-phosphoramidite body, as an example, synthesis and purification of a polyribonucleotide chain, confirmation of a base sequence, and third polyribonucleotide chain using a polyribonucleotide chain However, the present invention is not limited to this.

Reference Example Synthesis of nucleoside 3'-0-phosphoramidite Nucleoside 3'-0-phosphoramidite was synthesized by a reaction represented by the following formula. : (In the formula, Thp represents a tetrahydropyranyl group, Tr represents a trityl group, B ′ represents any one of benzoyladenine, isobutyrylguanine, benzoylcytosine, and anisoyluracil.) Hereinafter, the compound (1) The synthesis when B 'is benzoyladenine, isobutyrylguanine, benzoylcytosine and anisoyluracil is described.

1) B '= benzoyl adenine benzoyl adenosine 2'-tetrahydropyranyl-5'-dimethoxytrityl ((1) 0.61 mg, 0.8 mmol
l) was dehydrated under pyridine azeotrope, and dichloromethane was added (2 ml)
And isopropylethylamine (0.56 ml, 3.2 mmol), and the mixture was replaced with nitrogen, and chlorodiisopropylaminomethylphosphine (0.19 ml, 0.96 mmol) was added dropwise over 2 minutes. After stirring at room temperature for 40 minutes to confirm that the raw materials had disappeared, ethyl acetate (30 ml) was added, and the mixture was washed with a saturated sodium bicarbonate solution and saturated saline. 1PS (manufactured by Whatman)
After filtration, the solution is distilled off and silica gel chromatography (Wa
kogel C-300, 15 g, ethyl acetate) to give the desired product as a foam.

Yield 0.72 (0.78 mmol) 98% ¹ H-NMR (CDCl ₃ ) δ ppm: 8.9 (brs, 1H, NH), 8.72 (s, 1H, H-8), 8.23 (s, 1H, H-)
2), 8.0 (m, 2H, o-Ar), 7.6-7.4 (m, 12H, m, p-Ar and
trityl), 6.8 (m, 4H, 2,2 ', 6,6'H of trityl), 6.30
(Dd, 1H, H-1'J _{1 ', 2'} = 5.7Hz), 5.1 (m, 1H, H-
2 '), 4.6 (m, 2H, aceta1H of Thp and H-3'), 4.4
(M, 1H, H-4 '), 4.0-3.4 (m, 6H, H-5',-OCH ₂ of
Thp and -NC H- ), 3.77 (s, 6H, -OCH ₃ of trityl), 3.
5-3.2 (dd, 3H, P- OC H 3 J p -OC H 3 = 13.0and 13.2H
z), 2.0-1.4 (m, 6H , -CH 2 -of Thp), 1.2-0.9 (m, 12
_{. H, -NCH (C H 3} ) 2 31 P-NMR (CDCl 3) δppm: 148.15,148.68 Compound (2) in B 'is isobutyryl guanine,
The compounds benzoylcytosine and anisoyluracil were also synthesized in a similar manner. The yield and physical properties are shown below.

2) B '= isobutylguanine, yield 88% ¹ H-NMR (CDCl ₃ ) δ ppm: 11.5 (m, 1 H, NH), 8.7 (m, 1 H, NH), 7.79 (s, 1 H, H-8),
7.7−7.2 (m, 9H, Ar of trityl), 6.8 (m, 4H, 2,2 ′, 6,
6'H of trityl), 5.9 (m, 2H, H-1'J _{1 ', 2'} = 7.3H
z and H-2 '), 4.8-4.0 (m, 3H, H-3', 4 'and ace
ta1H of Thp), 3.77 (s, 6H, -OCH ₃ of trityl), 3.7-
2.9 (m, 10H, H- 5 ', - OCH 2 - of Thp, -NC H -, P-OC
_{H 3, -COC H (CH 3} ) 2), 2.0-1.6 (m, 6H, -CH 2 - of Th
p), 1.4 (m, 12H , -NCH (C H 3) 2, 1.0-1.8 (m, 6H, -C
^{_{. OCH (CH 3) 2 31}} P-NMR (CDCl 3) δppm: 148.21,148.48 3) B '= benzoyl cytosine, yield ^{93% 1 H-NMR (CDCl} 3) δppm: 8.63 (brs, 1H, NH), 8.52 (d, 1H, H-5 J _5.6 = 7.1 Hz), 7.
9 (m, 2H, o-Ar), 7.7-7.3 (m, 13H, H-6, m, p-Ar and t
rityl), 6.9 (m, 4H, 2, 2 ', 6, 6'H of trityl), 6.1 (m,
1H, H-1 '), 5.3 (m, 1H, aceta1H of Thp), 4.7-4.0
(M, 3H, H-2 ', 3', 4 '), 3.83 (s, 6H, -OCH ₃ of trit
yl), 3.8-3.5 (m, 5H , H-5 ', - NC H - and OCH 2 of
Thp), 3.4 (dd, 3H , P-OCH 3, J P OC H 3 = 13.2 and 13.2H
z), 2.0-1.4 (m, 6H , -CH 2 - of Thp), 1.3-1.0 (m, 12
_{H, -NCH (C H 3)} 2). ³¹ P-NMR (CDCl ₃ ) δ ppm: 147.20, 147.94 4) B ′ = Anisoyluracil, yield: 94% ¹ H-NMR (CDCl ₃ ) δ ppm: 8.01 (d, 1H, H-6 J _5.6 = 8.3 Hz) ), 7.9 (m, 3H, H-6, o-A
r), 7.3 (m, 11H, m-Arand trityl), 6.9 (m, 4H, 2, 2 ',
6,6'H of trityl), 6.17 (m, 1H, H-1 '), 5.31 (d, 1
H, H-5, J = _5.6 = 8.3Hz), 5.0 (brs, 1H, aceta1H of Th
p), 4.6 (m, 2H, H-2 ', 3'), 4.3 (m, 1H, H-4 '), 3.
86 (s, 3H, -OCH ₃ of anisoyl), 3.81 (s, 6H, -OCH ₃ of anisoyl)
trityl), 3.5 (m, 5H, H-5 ',-NC H- and -OCH ₂ of
Thp), 3.3 (dd, 3H , P-OCH 3, J P -OC H 3 = 12.9 and 1
3.4Hz), 1.9-1.4 (m, 6H , -CH 2 - of Thp), 1.2-1.0
_{(M, 12H, -NCH (C} H 3) 2). ³¹ P-NMR (CDCl ₃ ) δ ppm: 147.80 Example 1. Synthesis of polyribonucleotide chain A chain (hereinafter abbreviated as IA) and B chain (hereinafter IB) of formula (I)
Abbreviated. ), C chain (hereinafter abbreviated as IC), A of formula (II)
A chain (hereinafter abbreviated as IIA) and a B chain of the formula (III) (hereinafter abbreviated as IIIB) were synthesized. That is, a nucleoside resin in which a 2′-hydroxyl group is protected with a tetrahydropyranyl group and a 5′-hydroxyl group is protected with a dimethoxytrityl group is acid-treated,
It was synthesized by the solid phase phosphoramidite method extending the chain in the 5'-direction. The method of synthesizing IA is described, but other polyribonucleotide chains were synthesized in the same manner.

 Chain elongation was performed according to the following formula.

(The symbols in the formula represent the following.

DMTr: ^bz A: Benzoyl adenine n: Number of condensed nucleotides: controlled pore gl
ass) Thp and B 'have the same meanings as described above. ).

An Applied Biosystems (ABI) model 380 DNA synthesizer was used as a polyribonucleotide chain synthesizer. Nucleoside 3'-0-phosphoramidite ((4) 18 mg, about 20 μmol) prepared so as to be dissolved in 0.16 ml of acetonitrile was used in bottle # 1- of a synthesizer.
Set to 4. 1% dichloroacetic acid / dichloromethane was set in the bolt # 1-4. The other reagents were from ABI.

First, the method of Kierzek et al. (Biochemistry, 25 , 7,840-7,
846 (1986)), 1% dichloroacetic acid / dichloromethane was allowed to act on the synthesized nucleoside resin ((3), 1 μmol) for 120 seconds to remove the dimethoxytrityl group. Next, after condensing for 120 seconds using nucleoside 3'-0-phosphoramidite ((4), 20 equivalents) and tetrazole (50 equivalents), a solution of DMAP (dimethylaminopyridine) -acetic anhydride-lutidine / THF was used. Phosphorus in the phosphoramidite was oxidized to phosphoric acid. By repeating the above reaction sequentially, polyribonucleotide (5) having the target base sequence was obtained.

Example 2. Removal of a protecting group in a polyribonucleotide chain and cleavage from a resin Removal of a protecting group in a polyribonucleotide chain and cleavage from a resin were performed according to the following formula: (The abbreviations in the formula represent the following.

B: Any one of adenine nucleotide, guanine nucleotide, cytosine nucleotide, and uracil nucleotide.

DMTr, Thp, n and B 'have the same meanings as described above. That is, a thiophenol-triethylamine / dioxane solution was allowed to act on the compound (5) for 30 minutes to remove the methyl group of the phosphate-protecting group. Thereafter, the mixture was treated with a concentrated ammonium hydroxide solution at room temperature for 1 hour to cut out from the resin. The ammonium solution containing the excised polyribonucleotide was sealed and heated at 60 ° C. for 5 hours. Thereafter, the solvent was distilled off, and reversed phase column chromatography (C18, φ0.7 × 14 cm) was performed. A peak having a color of dimethoxytrityl cation due to perchloric acid was examined, and the fraction was collected and the solvent was distilled off. To this was added 0.01N hydrochloric acid, the pH was adjusted to 2.0 with 0.1N hydrochloric acid, and left at room temperature for 20 hours to remove the dimethoxytrityl group and the tetrahydropyranyl group. The mixture was neutralized with 0.1N ammonium hydroxide, washed with ethyl acetate, and then desalted with Sephadex G-25 (φ1.8 × 38 cm) to obtain a crude compound (6).

Example 3 Purification of Synthetic Polyribonucleotide Chain The crude compound (6) obtained in Example 2 was purified by reversed phase high performance liquid chromatography and ion exchange high performance liquid chromatography.

For reversed-phase high-performance liquid chromatography, see YMC
PACK C18 column (φ1.0 × 30cm, manufactured by Yamamura Science Co., Ltd.)
This was performed using Purification was performed by a 10-15% acetonitrile concentration gradient method using a 0.1 M triethylammonium acetate buffer (flow rate: 2 ml / min.). The RNA fraction was collected at an absorption wavelength of 254 nm. FIG. 1 shows the separation pattern of reversed-phase high performance liquid chromatography. The horizontal axis represents time, and the vertical axis represents the absorption at 254 nm and the concentration of acetonitrile.

The ion exchange chromatography was performed using a TSK gel DEAE-2SW column (φ0.46 × 25 cm, manufactured by Toyo Soda Co., Ltd.). Using 20% acetonitrile, 0.5-0.9 M ammonium formate concentration gradient method,
Purification was performed (flow rate Iml / min.). The polyribonucleotide fraction was collected with an absorption wavelength of 254 mm. FIG. 2 shows the separation pattern of ion exchange high performance liquid chromatography. The horizontal axis represents time, and the vertical axis represents absorption at 254 nm and the concentration of ammonium formate.

By the above separation and purification operations, IA of 4.75A and ₂₆₀ units was obtained.

Example 4 Confirmation of Base Sequence of Synthetic Polyribonucleotide Chain IA (0.015A ₂₆₀ units, 100 pmol) contains [γ- ³² P] ATP
(0.5 μl), buffer (1 μl, 250 mM Tris-HC1 (pH 7.
6), 50 mM magnesium chloride, 50 mM mercaptoethanol), T4 polynucleotide kinase (0.5 μl, 1 unit) and sterilized water (3 μl) were added, and the mixture was kept at 37 ° C. for 1 hour. To remove unreacted [γ- ³² P] ATP, spot on DEAE cellulose TLC and use Homomix III (Nucleic Acid Res.
earch, 1 , 331-353 (1974)). Remove the radioactive portion containing polyribonucleotides and use DEAE
Eluted from cellulose with 2M triethylammonium bicarbonate.

Add snake venom phosdiesterase (0.5μl, 1mg / 0.5ml)
Was added, and the mixture was kept at 37 ° C., and fractionated 30 minutes later to obtain a limited hydrolyzate. For this, electrophoresis at pH 3.5 was performed in the first dimension and homochromatography was performed in the second dimension to confirm the sequence.

The results for IA, IB and IC are shown in FIG.
4 and 5. 3, 4 and 5, the horizontal axis indicates the electrophoresis migration direction, and the vertical axis indicates the homochromatography migration direction.

Confirmation of chain length of synthetic polyribonucleotide The 5 'end of polyribonucleotides IIA and IIIB was labeled, and the chain length was confirmed by performing 20% polyacrylamide gel electrophoresis containing 8M urea. FIG. 6 shows the result.
(In the figure, lane l: IA, lane 2: IB, lane 3: IC, lane 4:
IIA, lane 5: IIIB, XC in the figure indicates xylene cyanol. Example 5 Cleavage reaction of another polyribonucleotide chain by two polyribonucleotide chains The IC used as a substrate was 5'-terminal [γ- ³² P] ATP, T4
After labeling with polyribonucleotide kinase, NENSORB 20
Desalinated and deproteinized by [Dopont] was used.

The cleavage reaction was performed as follows. 2.5 pmol of IC,
And IA and IB corresponding to catalytic polyribonucleotides
(Lane 3), II A and II B (lane 4) or IA and III B
(Lane 5) in 3.75 pmol each of 25 mM magnesium chloride, 40 mM Tris-HCl (pH 7.5), 20 mM salt solution (2.3
μl) and kept at 15 ° C. for 15 hours. Lane as a control
A buffer solution containing 5 mM EDTA instead of 25 mM magnesium chloride in 3 was added and kept warm (lane 2). 50mM EDTA (3μ
l) was added to stop the reaction, and the mixture was analyzed by 20% polyacrylamide gel electrophoresis containing 8 M urea. Marker (lane 1)
It was electrophoresed those cutting the labeled IC 5 'end with RNase U _2. The cleavage ratio was determined by cutting out the gel and quantifying it with a liquid scintillation counter. Table 1 shows the results.

FIG. 7 shows the results of the electrophoresis. In FIG. 7, RNase U
₂ cleaves the bond between the 3′-position of the purine of the polyribonucleotide chain and the phosphate of the subsequent nucleotide, and A ₂ , G ₃ , A ₆ , A ₇ , and A ₈ used as markers are each 2nd adenine nucleotide from the 5 'side of IC, 3
The mobility of the fragment cleaved after the 6th, 7th, or 8th adenine nucleotide is shown. Therefore, other lanes indicate that the cleavage was performed after the sixth adenine nucleotide.

[Brief description of the drawings]

FIG. 1 shows a separation pattern of a synthetic polyribonucleotide crude compound (IA) by reversed-phase high performance liquid chromatography. FIG. 2 shows a subsequent separation pattern by ion exchange high performance liquid chromatography. FIG. 3 shows patterns of IA, FIG. 4 shows IB, and FIG. 5 shows patterns of base sequence analysis by electrophoresis and homochromatography of IC. FIG. 6 shows a comparison pattern of chain lengths of IA, IB, IC, IIA and IIIB by electrophoresis. FIG. 7 shows an electrophoresis pattern of a fragment of IC by IA and IB, IIA and IB, or IA and IIIB.

Claims (3)

(57) [Claims] 1. A polynucleotide comprising a nucleotide sequence represented by A in the following formula (b) and excluding the nucleotide sequence represented by C, and a polynucleotide sequence comprising a nucleotide sequence represented by B and represented by C Method of cleaving a polyribonucleotide containing a nucleotide sequence represented by C at a site indicated by an arrow in the formula using a polyribonucleotide not containing: (In the formula, U represents a uracil nucleotide, A represents an adenine nucleotide, C represents a cytosine nucleotide, G represents a guanine nucleotide, R represents any of a uracil nucleotide, an adenine nucleotide, and a cytosine nucleotide, and corresponds to S _{3 to} S ₅ . Y ₁ corresponding to X _{3 to} X ₅ , V _{1 to} V ₂
~ Y ₂ , and Z _{3 to} Z ₅ corresponding to W _{3 to} W ₅ are uracil nucleotides, adenine nucleotides, which can form complementary hydrogen bonds, respectively, of which the integer numbers in the combination of these three groups match. cytosine nucleotides, represent either guanine nucleotide, W ₁ corresponding to _{X 1 ~X 2, V 3 ~V} 4 and the corresponding Y ₃ to Y _4, and Z ₁ to Z ₂ and the corresponding S ₁ to S ₂ ~ W ₂ is a uracil nucleotide, an adenine nucleotide, a cytosine nucleotide, in which at least two of the combinations of the three groups can form complementary hydrogen bonds, each having the same integer number,
Represents any of the guanine nucleotides. ). 2. The method according to claim 1, wherein S ₁ -S ₂
Corresponding to the _{X 1 ~X 2, V 3 ~V} 4 and the corresponding Y ₃ to Y _4, and Z ₁ ~
Any of uracil nucleotides, adenine nucleotides, cytosine nucleotides, and guanine nucleotides in which W _{1 to} W ₂ corresponding to Z ₂ are a combination of these three groups, and whose integer numbers match each other can form complementary hydrogen bonds. The method for cleaving a polyribonucleotide according to claim 1, wherein 3. The method according to claim 1, wherein R,
S ₁ , W ₁ , W ₃ , X ₅ , Y ₃ , Z ₂ and Z ₅ are adenine nucleotides, S ₂ , S ₃ , S ₄ , V ₂ , W ₄ , Y ₁ and Y ₄ are cytosine nucleotides, V ₁ , V ₄ , X ₂ , X ₃ , X ₄ , Y ₂ and Z ₄ are guanine nucleotides, and S ₅ , V ₃ , W ₂ , W ₅ , X ₁ , Z ₁ and Z ₃ are uracil nucleotides The method for cleaving a polyribonucleotide according to claim 1 or 2.