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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

Quantity of mRNA extracted from total RNA by SPGE (—) and number of copies of ACT7 after multiple usage of SPGE probes (- - -). The probes were washed before release of mRNA (80 °C for 3 min). Average ± SE, n = 9. Least significance difference (p < 0.05) for amounts of SPGE-extracted mRNA amounts of SPGE-extracted mRNA is 0.39 ng, and for number of copies of ACT7 is 1,403 copies
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Fig4: Quantity of mRNA extracted from total RNA by SPGE (—) and number of copies of ACT7 after multiple usage of SPGE probes (- - -). The probes were washed before release of mRNA (80 °C for 3 min). Average ± SE, n = 9. Least significance difference (p < 0.05) for amounts of SPGE-extracted mRNA amounts of SPGE-extracted mRNA is 0.39 ng, and for number of copies of ACT7 is 1,403 copies

Mentions: After washing and incubation in 80 °C DEPC-treated water for 3 min, each needle released up to 9.1 ng mRNA. However, repeated extractions with the same probes resulted in a drastic decrease of ACT7 quantification (Fig. 4).Fig. 4


Profiling Gene Expression in Germinating Brassica Roots.

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

Quantity of mRNA extracted from total RNA by SPGE (—) and number of copies of ACT7 after multiple usage of SPGE probes (- - -). The probes were washed before release of mRNA (80 °C for 3 min). Average ± SE, n = 9. Least significance difference (p < 0.05) for amounts of SPGE-extracted mRNA amounts of SPGE-extracted mRNA is 0.39 ng, and for number of copies of ACT7 is 1,403 copies
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3926982&req=5

Fig4: Quantity of mRNA extracted from total RNA by SPGE (—) and number of copies of ACT7 after multiple usage of SPGE probes (- - -). The probes were washed before release of mRNA (80 °C for 3 min). Average ± SE, n = 9. Least significance difference (p < 0.05) for amounts of SPGE-extracted mRNA amounts of SPGE-extracted mRNA is 0.39 ng, and for number of copies of ACT7 is 1,403 copies
Mentions: After washing and incubation in 80 °C DEPC-treated water for 3 min, each needle released up to 9.1 ng mRNA. However, repeated extractions with the same probes resulted in a drastic decrease of ACT7 quantification (Fig. 4).Fig. 4

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