Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification
Strand displacement amplification (SDA) is an isothermal amplification technique wherein amplification of a nucleic acid is initiated by nicking enzyme activity at sites flanking the target. Diagnostic SDA is very fast but requires precise optimization and is limited to very short amplicons. Here we...
Saved in:
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Taylor & Francis Group
2024-06-01
|
| Series: | BioTechniques |
| Subjects: | |
| Online Access: | https://www.tandfonline.com/doi/10.2144/btn-2024-0012 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1841526670932049920 |
|---|---|
| author | Yinhua Zhang Ashley N Luck Nathan A Tanner |
| author_facet | Yinhua Zhang Ashley N Luck Nathan A Tanner |
| author_sort | Yinhua Zhang |
| collection | DOAJ |
| description | Strand displacement amplification (SDA) is an isothermal amplification technique wherein amplification of a nucleic acid is initiated by nicking enzyme activity at sites flanking the target. Diagnostic SDA is very fast but requires precise optimization and is limited to very short amplicons. Here we report an enhanced approach by addition of single-stranded DNA binding protein, crowding agents and dUTP to enable amplification of kilobase-length products at low temperatures. Additionally, we pair this improved SDA with a novel carryover contamination prevention, eliminating amplifiable DNA at the end of the reaction to reduce contamination risk. Taken together these developments increase the utility and versatility of SDA, broadening the reach of this powerful but uncommonly used method. |
| format | Article |
| id | doaj-art-a75311cd27984302807e023c6810bf7e |
| institution | Kabale University |
| issn | 0736-6205 1940-9818 |
| language | English |
| publishDate | 2024-06-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | BioTechniques |
| spelling | doaj-art-a75311cd27984302807e023c6810bf7e2025-01-16T15:38:47ZengTaylor & Francis GroupBioTechniques0736-62051940-98182024-06-0176625526210.2144/btn-2024-0012Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplificationYinhua Zhang0Ashley N Luck1Nathan A Tanner2New England Biolabs, 240 County Road, Ipswich, MA 01938, USANew England Biolabs, 240 County Road, Ipswich, MA 01938, USANew England Biolabs, 240 County Road, Ipswich, MA 01938, USAStrand displacement amplification (SDA) is an isothermal amplification technique wherein amplification of a nucleic acid is initiated by nicking enzyme activity at sites flanking the target. Diagnostic SDA is very fast but requires precise optimization and is limited to very short amplicons. Here we report an enhanced approach by addition of single-stranded DNA binding protein, crowding agents and dUTP to enable amplification of kilobase-length products at low temperatures. Additionally, we pair this improved SDA with a novel carryover contamination prevention, eliminating amplifiable DNA at the end of the reaction to reduce contamination risk. Taken together these developments increase the utility and versatility of SDA, broadening the reach of this powerful but uncommonly used method.https://www.tandfonline.com/doi/10.2144/btn-2024-0012diagnosticsDNA amplificationisothermal amplification |
| spellingShingle | Yinhua Zhang Ashley N Luck Nathan A Tanner Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification BioTechniques diagnostics DNA amplification isothermal amplification |
| title | Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification |
| title_full | Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification |
| title_fullStr | Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification |
| title_full_unstemmed | Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification |
| title_short | Isothermal amplification of long DNA fragments at low temperature by improved strand displacement amplification |
| title_sort | isothermal amplification of long dna fragments at low temperature by improved strand displacement amplification |
| topic | diagnostics DNA amplification isothermal amplification |
| url | https://www.tandfonline.com/doi/10.2144/btn-2024-0012 |
| work_keys_str_mv | AT yinhuazhang isothermalamplificationoflongdnafragmentsatlowtemperaturebyimprovedstranddisplacementamplification AT ashleynluck isothermalamplificationoflongdnafragmentsatlowtemperaturebyimprovedstranddisplacementamplification AT nathanatanner isothermalamplificationoflongdnafragmentsatlowtemperaturebyimprovedstranddisplacementamplification |