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Published on: August 2, 2014
Comparison of amylase mRNAs from rat parotid gland, pancreas and liver using reverse transcriptase-polymerase chain
Akihiro Nezu1, Takao Morita, Akihiko Tanimura
1Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan. a-nezu@hoku-iryo-u-ac.jp
Abstract:
The expression of mRNA for amylase was examined using the reverse transcriptase-polymerase chain reaction (RT-PCR). An amylase product was strongly detected in parotid and pancreas, but less strongly in liver. The degree of identity between the PCR products was assessed by restriction-enzyme mapping using two restriction enzymes, EcoRI and ScaI, and DNA sequencing. The PCR product from pancreas was cut by both EcoRI and ScaI, while the products from parotid and liver were cut by EcoRI but not by ScaI. The sequence of the parotid product was 90.4% homologous to that of the pancreas, and 100% homologous to that of the liver. These results indicate that the same amylase mRNA may be expressed in parotid and liver. In addition, the expression of amylase mRNAs in other rat tissues was investigated using RT-PCR, and the sensitivity of each PCR product to ScaI was tested. A weak single band was detected in submandibular gland, sublingual gland and stomach. ScaI digestion cut the stomach product into two fragments, but had no effect on the submandibular and sublingual products. Thus, it may be possible to classify amylase isoenzymes into pancreatic and parotid types based on the sensitivity of their PCR products to ScaI.
Insights
Researchers identified distinct amylase isoenzymes in rats using reverse transcriptase-polymerase chain reaction (RT-PCR). Amylase mRNA expression varies across tissues, suggesting different amylase types in pancreas and parotid glands.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Amylase is a crucial enzyme for carbohydrate digestion.
- Different tissues express amylase, but the specific isoenzymes and their regulation are not fully understood.
Purpose of the Study:
- To investigate the expression of amylase mRNA in various rat tissues.
- To characterize and differentiate amylase isoenzymes based on molecular properties.
Main Methods:
- Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect amylase mRNA.
- Restriction enzyme mapping with EcoRI and ScaI, and DNA sequencing were employed to analyze PCR products.
- Amylase mRNA expression was analyzed in parotid, pancreas, liver, submandibular gland, sublingual gland, and stomach tissues.
Main Results:
- Amylase mRNA was strongly detected in parotid and pancreas, and less so in the liver.
- PCR products from pancreas were sensitive to both EcoRI and ScaI, while parotid and liver products were only sensitive to EcoRI.
- Parotid amylase mRNA showed 90.4% homology to pancreatic and 100% to liver amylase mRNA, suggesting shared expression.
- Amylase mRNA was also detected in submandibular gland, sublingual gland, and stomach, with distinct ScaI sensitivity patterns.
Conclusions:
- The findings suggest the expression of similar amylase mRNA in parotid and liver tissues.
- Amylase isoenzymes can be classified into pancreatic and parotid types based on the ScaI enzyme sensitivity of their respective PCR products.
