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Cloning of a complementary deoxyribonucleic acid encoding a portion of rat intestinal preapolipoprotein AIV messenger
Abstract:
Apolipoprotein AIV is one of the principal apolipoproteins synthesized by the rat small intestine. We have cloned a cDNA encoding a portion of preapolipoprotein AIV mRNA. A kinetically fractionated cDNA probe highly enriched for the abundant intestinal mucosal mRNA sequences was used to screen a library of recombinants containing cDNA generated from total intestinal epithelial mRNA. The abundant class of mRNA sequences was defined by hybridization analyses. This frequency class had an aggregate complexity of 5300 nucleotides and represented 25% of accumulated mRNA sequences.l The mRNAs comprising this class were identified by in vitro translation and included preapolipoprotein AIV, preproapolipoprotein AI, intestinal fatty acid binding protein, and liver fatty acid binding protein. A cDNA-containing clone derived from preapolipoprotein AIV mRNA was identified among probe-positive recombinants. This cDNA was used to establish that apolipoprotein AIV mRNA has a mass of 550000 daltons (equivalent to 1780 nucleotides) and represents 0.013% of total cellular RNA in the fasting state. Acute feeding with triglyceride-rich meals resulted in a 2-fold increase in preapolipoprotein AiV mRNA after 4 h.
Insights
Researchers cloned a complementary DNA (cDNA) for rat preapolipoprotein AIV (ApoAIV) mRNA from the small intestine. Dietary fat intake significantly increased ApoAIV mRNA levels, suggesting its role in lipid absorption.
Area of Science:
- Molecular Biology
- Gastroenterology
- Biochemistry
Background:
- Apolipoprotein AIV (ApoAIV) is a key protein synthesized in the rat small intestine.
- Understanding ApoAIV synthesis regulation is crucial for comprehending lipid metabolism.
Purpose of the Study:
- To clone the complementary DNA (cDNA) encoding rat preapolipoprotein AIV (preApoAIV) mRNA.
- To investigate the regulation of preApoAIV mRNA levels in response to dietary stimuli.
Main Methods:
- Screening a rat intestinal epithelial cell cDNA library using a kinetically fractionated probe.
- Hybridization analyses to define abundant mRNA frequency classes.
- In vitro translation to identify specific mRNAs.
- Northern blot analysis to determine mRNA mass and abundance.
Main Results:
- A cDNA clone for preApoAIV mRNA was successfully isolated.
- ApoAIV mRNA has a mass of 1780 nucleotides and constitutes 0.013% of total cellular RNA in fasting rats.
- Feeding triglyceride-rich meals doubled preApoAIV mRNA levels within 4 hours.
Conclusions:
- The study successfully cloned preApoAIV cDNA and characterized its mRNA.
- Dietary fat intake significantly upregulates preApoAIV mRNA expression in the rat small intestine.
- These findings highlight ApoAIV's role in intestinal lipid processing and transport.