METHODS FOR AMPLIFYING NUCLEIC ACIDS AND FOR ANALYZING NUCLEIC ACIDS THEREWITH
First Claim
1. A method for amplifying a nucleic acid, comprising:
- a complementary strand amplifying step, in which a target double-stranded nucleic acid to be amplified, and a first primer complementary to a region in one strand of said nucleic acid are used;
a second-primer adding step, in which a second primer complementary to a region on the 3′
side of the amplified product from said complementary strand amplifying step is added;
a double strand amplifying step in which said target double-stranded nucleic acid to be amplified is amplified in the presence of the first primer and the second primer.
1 Assignment
0 Petitions
Accused Products
Abstract
An object of the present invention is to control the increase in amplification errors generated during the nucleic acid amplification and thus to obtain an amplification product having a good reproducibility. The present invention is characterized in that a target nucleic acid to be amplified is amplified through a two-stage amplification process in which the amplification only of a single strand is first performed and a strand which is complementary to its amplified product is then amplified. The amplification uses a first primer which is employed for the first-stage amplification and a second primer which is employed for the second-stage amplification. These primers are each used separately, or alternatively designed to have a different stringency and used at the same time.
-
Citations
14 Claims
-
1. A method for amplifying a nucleic acid, comprising:
-
a complementary strand amplifying step, in which a target double-stranded nucleic acid to be amplified, and a first primer complementary to a region in one strand of said nucleic acid are used; a second-primer adding step, in which a second primer complementary to a region on the 3′
side of the amplified product from said complementary strand amplifying step is added;a double strand amplifying step in which said target double-stranded nucleic acid to be amplified is amplified in the presence of the first primer and the second primer. - View Dependent Claims (7, 8, 9, 10, 11, 12, 13, 14)
-
-
2. A method for amplifying a nucleic acid, comprising:
-
a complementary strand amplifying step, in which a target single-stranded nucleic acid to be amplified, and a first primer complementary to a region in said nucleic acid, are used; a second-primer adding step, in which a second primer complementary to a region on the 3′
side of the amplified product from said complementary strand amplifying step is added; anda double strand amplifying step, in which said target nucleic acid to be amplified is amplified in the presence of the first primer and the second primer.
-
-
3. A method for amplifying a nucleic acid, comprising:
-
an amplification preparing step, in which a target double-stranded nucleic acid to be amplified, a first primer complementary to a region in one strand of said nucleic acid, and a second primer complementary to a region in the other strand of said nucleic acid and optimal stringency of which being significantly milder than that of the first primer are mixed; a first amplification step, being carried out under conditions having an optimal stringency for the combination of the first primer and the target nucleic acid to be amplified; and a second amplification step, being carried out under conditions having an optimal stringency for the combination of the second primer and the target nucleic acid to be amplified. - View Dependent Claims (5, 6)
-
-
4. A method for amplifying a nucleic acid, comprising:
-
an amplification preparing step, in which a target single-stranded nucleic acid to be amplified, a first primer complementary to a region in said nucleic acid, and a second primer complementary to a region on the 3′
side of the extension product extended with said first primer and optimal stringency of which being significantly milder than that of said first primer are mixed;a first amplification step, being carried out under conditions having an optimal stringency for the combination of the first primer and the target nucleic acid to be amplified; and a second amplification step, being carried out under conditions having an optimal stringency for the combination of the second primer and the amplification product of the first amplification step.
-
Specification