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Profiling Gene Expression in Germinating Brassica Roots.

Park MR, Wang YH, Hasenstein KH - Plant Mol. Biol. Rep. (2014)

Bottom Line: The expression level of all genes differed significantly at each sample position.SPGE needles can be used within two weeks when stored at 4 °C.Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

View Article: PubMed Central - PubMed

ABSTRACT
Based on previously developed solid-phase gene extraction (SPGE) we examined the mRNA profile in primary roots of Brassica rapa seedlings for highly expressed genes like ACT7 (actin7), TUB (tubulin1), UBQ (ubiquitin), and low expressed GLK (glucokinase) during the first day post-germination. The assessment was based on the mRNA load of the SPGE probe of about 2.1 ng. The number of copies of the investigated genes changed spatially along the length of primary roots. The expression level of all genes differed significantly at each sample position. Among the examined genes ACT7 expression was most even along the root. UBQ was highest at the tip and root-shoot junction (RS). TUB and GLK showed a basipetal gradient. The temporal expression of UBQ was highest in the MZ 9 h after primary root emergence and higher than at any other sample position. Expressions of GLK in EZ and RS increased gradually over time. SPGE extraction is the result of oligo-dT and oligo-dA hybridization and the results illustrate that SPGE can be used for gene expression profiling at high spatial and temporal resolution. SPGE needles can be used within two weeks when stored at 4 °C. Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

No MeSH data available.


Related in: MedlinePlus

Expression of ubiquitin (UBQ, top) and glucokinase (GLK, bottom) of Brasscia primary roots. The roots were sampled at the meristem (square), elongation zone (diamond) and root-shoot junction (circle) between 6 and 24 h after germination; average ± SE, n = 9. Least significance differences (p < 0.05) for UBQ copy numbers in MZ, EZ, and RS are 164, 683, and 666 copies, respectively; for GLK in MZ, EZ, and RS, the values are 5.7, 18.7, and 10.8 copies, respectively
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Fig7: Expression of ubiquitin (UBQ, top) and glucokinase (GLK, bottom) of Brasscia primary roots. The roots were sampled at the meristem (square), elongation zone (diamond) and root-shoot junction (circle) between 6 and 24 h after germination; average ± SE, n = 9. Least significance differences (p < 0.05) for UBQ copy numbers in MZ, EZ, and RS are 164, 683, and 666 copies, respectively; for GLK in MZ, EZ, and RS, the values are 5.7, 18.7, and 10.8 copies, respectively

Mentions: We used two genes of different expression levels (UBQ, high expression and GLK, low expression), to investigate the dynamic range and profiling of genes over time at three positions (MZ, EZ, and RS; Fig. 7). The expression of these two genes showed clearly varied during the first 24 h after primary root emergence. SPGE covers a dynamic range of eightfold at the EZ after 24 h for UBQ. The dynamic range increased to 107-fold at RS after 24 h for UBQ compared to that of GLK. Similar to positional changes, the expression of all examined genes changed over time. The copy number of UBQ fluctuated but remained generally low in the MZ and EZ but expression at the RS region strongly increased after 18 h. In contrast, the expression of GLK showed a steady increase at all sampling times for RS and EZ but remained low in the MZ (Fig. 7). Therefore, the expression patterns of the examined genes and most likely other, if not most, genes vary over time and position. We verified that the differences in gene expressions are not the result of different extraction yields based on tissue or developmental stage. The yield per probe at different extraction sites was not different and averaged 2.1 ± 0.2 ng (data not shown).Fig. 7


Profiling Gene Expression in Germinating Brassica Roots.

Park MR, Wang YH, Hasenstein KH - Plant Mol. Biol. Rep. (2014)

Expression of ubiquitin (UBQ, top) and glucokinase (GLK, bottom) of Brasscia primary roots. The roots were sampled at the meristem (square), elongation zone (diamond) and root-shoot junction (circle) between 6 and 24 h after germination; average ± SE, n = 9. Least significance differences (p < 0.05) for UBQ copy numbers in MZ, EZ, and RS are 164, 683, and 666 copies, respectively; for GLK in MZ, EZ, and RS, the values are 5.7, 18.7, and 10.8 copies, respectively
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig7: Expression of ubiquitin (UBQ, top) and glucokinase (GLK, bottom) of Brasscia primary roots. The roots were sampled at the meristem (square), elongation zone (diamond) and root-shoot junction (circle) between 6 and 24 h after germination; average ± SE, n = 9. Least significance differences (p < 0.05) for UBQ copy numbers in MZ, EZ, and RS are 164, 683, and 666 copies, respectively; for GLK in MZ, EZ, and RS, the values are 5.7, 18.7, and 10.8 copies, respectively
Mentions: We used two genes of different expression levels (UBQ, high expression and GLK, low expression), to investigate the dynamic range and profiling of genes over time at three positions (MZ, EZ, and RS; Fig. 7). The expression of these two genes showed clearly varied during the first 24 h after primary root emergence. SPGE covers a dynamic range of eightfold at the EZ after 24 h for UBQ. The dynamic range increased to 107-fold at RS after 24 h for UBQ compared to that of GLK. Similar to positional changes, the expression of all examined genes changed over time. The copy number of UBQ fluctuated but remained generally low in the MZ and EZ but expression at the RS region strongly increased after 18 h. In contrast, the expression of GLK showed a steady increase at all sampling times for RS and EZ but remained low in the MZ (Fig. 7). Therefore, the expression patterns of the examined genes and most likely other, if not most, genes vary over time and position. We verified that the differences in gene expressions are not the result of different extraction yields based on tissue or developmental stage. The yield per probe at different extraction sites was not different and averaged 2.1 ± 0.2 ng (data not shown).Fig. 7

Bottom Line: The expression level of all genes differed significantly at each sample position.SPGE needles can be used within two weeks when stored at 4 °C.Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

View Article: PubMed Central - PubMed

ABSTRACT
Based on previously developed solid-phase gene extraction (SPGE) we examined the mRNA profile in primary roots of Brassica rapa seedlings for highly expressed genes like ACT7 (actin7), TUB (tubulin1), UBQ (ubiquitin), and low expressed GLK (glucokinase) during the first day post-germination. The assessment was based on the mRNA load of the SPGE probe of about 2.1 ng. The number of copies of the investigated genes changed spatially along the length of primary roots. The expression level of all genes differed significantly at each sample position. Among the examined genes ACT7 expression was most even along the root. UBQ was highest at the tip and root-shoot junction (RS). TUB and GLK showed a basipetal gradient. The temporal expression of UBQ was highest in the MZ 9 h after primary root emergence and higher than at any other sample position. Expressions of GLK in EZ and RS increased gradually over time. SPGE extraction is the result of oligo-dT and oligo-dA hybridization and the results illustrate that SPGE can be used for gene expression profiling at high spatial and temporal resolution. SPGE needles can be used within two weeks when stored at 4 °C. Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

No MeSH data available.


Related in: MedlinePlus