Stool sample storage conditions for the preservation of Giardia intestinalis DNA
Stool is chemically complex and the extraction of DNA from stool samples is extremely difficult. Haemoglobin breakdown products, such as bilirubin, bile acids and mineral ions, that are present in the stool samples, can inhibit DNA amplification and cause molecular assays to produce false-negative r...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Fundação Oswaldo Cruz (FIOCRUZ)
2012-12-01
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| Series: | Memorias do Instituto Oswaldo Cruz |
| Subjects: | |
| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000800001&lng=en&tlng=en |
| _version_ | 1797703094074605568 |
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| author | Salih Kuk Suleyman Yazar Ulfet Cetinkaya |
| author_facet | Salih Kuk Suleyman Yazar Ulfet Cetinkaya |
| author_sort | Salih Kuk |
| collection | DOAJ |
| description | Stool is chemically complex and the extraction of DNA from stool samples is extremely difficult. Haemoglobin breakdown products, such as bilirubin, bile acids and mineral ions, that are present in the stool samples, can inhibit DNA amplification and cause molecular assays to produce false-negative results. Therefore, stool storage conditions are highly important for the diagnosis of intestinal parasites and other microorganisms through molecular approaches. In the current study, stool samples that were positive for Giardia intestinalis were collected from five different patients. Each sample was stored using one out of six different storage conditions [room temperature (RT), +4ºC, -20ºC, 70% alcohol, 10% formaldehyde or 2.5% potassium dichromate] for DNA extraction procedures at one, two, three and four weeks. A modified QIAamp Stool Mini Kit procedure was used to isolate the DNA from stored samples. After DNA isolation, polymerase chain reaction (PCR) amplification was performed using primers that target the β-giardin gene. A G. intestinalis-specific 384 bp band was obtained from all of the cyst-containing stool samples that were stored at RT, +4ºC and -20ºC and in 70% alcohol and 2.5% potassium dichromate; however, this band was not produced by samples that had been stored in 10% formaldehyde. Moreover, for the stool samples containing trophozoites, the same G. intestinalis-specific band was only obtained from the samples that were stored in 2.5% potassium dichromate for up to one month. As a result, it appears evident that the most suitable storage condition for stool samples to permit the isolation of G. intestinalis DNA is in 2.5% potassium dichromate; under these conditions, stool samples may be stored for one month. |
| first_indexed | 2024-03-12T04:59:26Z |
| format | Article |
| id | doaj.art-27fc3798d0fc49dbb3e10caead75c3b6 |
| institution | Directory Open Access Journal |
| issn | 1678-8060 |
| language | English |
| last_indexed | 2024-03-12T04:59:26Z |
| publishDate | 2012-12-01 |
| publisher | Fundação Oswaldo Cruz (FIOCRUZ) |
| record_format | Article |
| series | Memorias do Instituto Oswaldo Cruz |
| spelling | doaj.art-27fc3798d0fc49dbb3e10caead75c3b62023-09-03T09:14:09ZengFundação Oswaldo Cruz (FIOCRUZ)Memorias do Instituto Oswaldo Cruz1678-80602012-12-01107896596810.1590/S0074-02762012000800001S0074-02762012000800001Stool sample storage conditions for the preservation of Giardia intestinalis DNASalih Kuk0Suleyman Yazar1Ulfet Cetinkaya2Erciyes UniversityErciyes UniversityErciyes UniversityStool is chemically complex and the extraction of DNA from stool samples is extremely difficult. Haemoglobin breakdown products, such as bilirubin, bile acids and mineral ions, that are present in the stool samples, can inhibit DNA amplification and cause molecular assays to produce false-negative results. Therefore, stool storage conditions are highly important for the diagnosis of intestinal parasites and other microorganisms through molecular approaches. In the current study, stool samples that were positive for Giardia intestinalis were collected from five different patients. Each sample was stored using one out of six different storage conditions [room temperature (RT), +4ºC, -20ºC, 70% alcohol, 10% formaldehyde or 2.5% potassium dichromate] for DNA extraction procedures at one, two, three and four weeks. A modified QIAamp Stool Mini Kit procedure was used to isolate the DNA from stored samples. After DNA isolation, polymerase chain reaction (PCR) amplification was performed using primers that target the β-giardin gene. A G. intestinalis-specific 384 bp band was obtained from all of the cyst-containing stool samples that were stored at RT, +4ºC and -20ºC and in 70% alcohol and 2.5% potassium dichromate; however, this band was not produced by samples that had been stored in 10% formaldehyde. Moreover, for the stool samples containing trophozoites, the same G. intestinalis-specific band was only obtained from the samples that were stored in 2.5% potassium dichromate for up to one month. As a result, it appears evident that the most suitable storage condition for stool samples to permit the isolation of G. intestinalis DNA is in 2.5% potassium dichromate; under these conditions, stool samples may be stored for one month.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000800001&lng=en&tlng=enGiardia intestinalisDNA isolationstorage conditionsstool |
| spellingShingle | Salih Kuk Suleyman Yazar Ulfet Cetinkaya Stool sample storage conditions for the preservation of Giardia intestinalis DNA Memorias do Instituto Oswaldo Cruz Giardia intestinalis DNA isolation storage conditions stool |
| title | Stool sample storage conditions for the preservation of Giardia intestinalis DNA |
| title_full | Stool sample storage conditions for the preservation of Giardia intestinalis DNA |
| title_fullStr | Stool sample storage conditions for the preservation of Giardia intestinalis DNA |
| title_full_unstemmed | Stool sample storage conditions for the preservation of Giardia intestinalis DNA |
| title_short | Stool sample storage conditions for the preservation of Giardia intestinalis DNA |
| title_sort | stool sample storage conditions for the preservation of giardia intestinalis dna |
| topic | Giardia intestinalis DNA isolation storage conditions stool |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000800001&lng=en&tlng=en |
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