Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it>
<p>Abstract</p> <p>Background</p> <p>Real-time RT-PCR has become a powerful technique to monitor low-abundance mRNA expression and is a useful tool when examining bacterial gene expression inside infected host tissues. However, correct evaluation of data requires accura...
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| Format: | Article |
| Language: | English |
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BMC
2007-09-01
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| Series: | BMC Plant Biology |
| Online Access: | http://www.biomedcentral.com/1471-2229/7/50 |
| _version_ | 1811249414806700032 |
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| author | Toth Ian K Takle Gunnhild W Brurberg May B |
| author_facet | Toth Ian K Takle Gunnhild W Brurberg May B |
| author_sort | Toth Ian K |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Real-time RT-PCR has become a powerful technique to monitor low-abundance mRNA expression and is a useful tool when examining bacterial gene expression inside infected host tissues. However, correct evaluation of data requires accurate and reliable normalisation against internal standards. Thus, the identification of reference genes whose expression does not change during the course of the experiment is of paramount importance. Here, we present a study where manipulation of cultural growth conditions and <it>in planta </it>experiments have been used to validate the expression stability of reference gene candidates for the plant pathogen <it>Pectobacterium atrosepticum</it>, belonging to the family <it>Enterobacteriaceae</it>.</p> <p>Results</p> <p>Of twelve reference gene candidates tested, four proved to be stably expressed both in six different cultural growth conditions and <it>in planta</it>. Two of these genes (<it>recA </it>and <it>ffh</it>), encoding recombinase A and signal recognition particle protein, respectively, proved to be the most stable set of reference genes under the experimental conditions used. In addition, genes <it>proC </it>and <it>gyrA</it>, encoding pyrroline-5-carboxylate reductase and DNA gyrase, respectively, also displayed relatively stable mRNA expression levels.</p> <p>Conclusion</p> <p>Based on these results, we suggest <it>recA </it>and <it>ffh </it>as suitable candidates for accurate normalisation of real-time RT-PCR data for experiments investigating the plant pathogen <it>P. atrosepticum </it>and potentially other related pathogens.</p> |
| first_indexed | 2024-04-12T15:46:07Z |
| format | Article |
| id | doaj.art-1344e178bf984a539e612fb537127f4d |
| institution | Directory Open Access Journal |
| issn | 1471-2229 |
| language | English |
| last_indexed | 2024-04-12T15:46:07Z |
| publishDate | 2007-09-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Plant Biology |
| spelling | doaj.art-1344e178bf984a539e612fb537127f4d2022-12-22T03:26:38ZengBMCBMC Plant Biology1471-22292007-09-01715010.1186/1471-2229-7-50Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it>Toth Ian KTakle Gunnhild WBrurberg May B<p>Abstract</p> <p>Background</p> <p>Real-time RT-PCR has become a powerful technique to monitor low-abundance mRNA expression and is a useful tool when examining bacterial gene expression inside infected host tissues. However, correct evaluation of data requires accurate and reliable normalisation against internal standards. Thus, the identification of reference genes whose expression does not change during the course of the experiment is of paramount importance. Here, we present a study where manipulation of cultural growth conditions and <it>in planta </it>experiments have been used to validate the expression stability of reference gene candidates for the plant pathogen <it>Pectobacterium atrosepticum</it>, belonging to the family <it>Enterobacteriaceae</it>.</p> <p>Results</p> <p>Of twelve reference gene candidates tested, four proved to be stably expressed both in six different cultural growth conditions and <it>in planta</it>. Two of these genes (<it>recA </it>and <it>ffh</it>), encoding recombinase A and signal recognition particle protein, respectively, proved to be the most stable set of reference genes under the experimental conditions used. In addition, genes <it>proC </it>and <it>gyrA</it>, encoding pyrroline-5-carboxylate reductase and DNA gyrase, respectively, also displayed relatively stable mRNA expression levels.</p> <p>Conclusion</p> <p>Based on these results, we suggest <it>recA </it>and <it>ffh </it>as suitable candidates for accurate normalisation of real-time RT-PCR data for experiments investigating the plant pathogen <it>P. atrosepticum </it>and potentially other related pathogens.</p>http://www.biomedcentral.com/1471-2229/7/50 |
| spellingShingle | Toth Ian K Takle Gunnhild W Brurberg May B Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it> BMC Plant Biology |
| title | Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it> |
| title_full | Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it> |
| title_fullStr | Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it> |
| title_full_unstemmed | Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it> |
| title_short | Evaluation of reference genes for real-time RT-PCR expression studies in the plant pathogen <it>Pectobacterium atrosepticum</it> |
| title_sort | evaluation of reference genes for real time rt pcr expression studies in the plant pathogen it pectobacterium atrosepticum it |
| url | http://www.biomedcentral.com/1471-2229/7/50 |
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