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GLOBEC: Physiological Status of Zooplankton: Utilizing PCR to Rapidly Assess Specific Enzyme Expression

GLOBEC：浮游动物的生理状态：利用 PCR 快速评估特定酶的表达

基本信息

批准号：
9116016
负责人：
Douglas Crawford
金额：
$ 33.96万
依托单位：
University of Chicago
依托单位国家：
美国
项目类别：
Continuing Grant
财政年份：
1992
资助国家：
美国
起止时间：
1992-01-15 至 1995-12-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=9116016&HistoricalAwards=false
关键词：
GLOBEC Physiological Status Zooplankton Utilizing

项目摘要

This project offers a unique solution to the problems associated with identifying, sorting and assessing the physiological status of zooplankton. Molecular techniques will be developed that can rapidly identify and quantify the level of expression of physiologically significant genes in the copepod, Calanus finmarchicus. These techniques take advantage of the specificity and sensitivity of the polymerase chain reaction (PCR) to quantitatively amplify species-specific mRNAs that encode enzymes that regulate metabolic flux. In order to design the necessary primers for PCR, the cDNAs that encode for the enzymes, hexokinase, phosphofructokinase, phosphoglucose isomerase and citrate synthetase will be cloned utilizing the phylogenetically conserved primary sequences of these enzymes. Species-specific primers will be designed from these cDNA sequences and used to quantitatively amplify each enzyme's mRNA only from C. finmarchicus to ascertain the affect of environmental parameters on the level of enzyme expression. Differences in the quantity of C. finmarchicus present in a taxonomically complex assemblage of zooplankton will be normalized by simultaneously measuring the specific concentration of a physiologically invariant gene, such as actin. The results will be equivalent to enzyme per weight of C. finmarchicus, a unit comparable to more traditional physiological studies.

该项目为相关问题提供了独特的解决方案通过识别、分类和评估浮游动物分子技术将被开发出来，快速鉴定和定量表达水平，桡足类哲水蚤的重要生理基因 finmarchicus。这些技术利用了聚合酶链反应（PCR）的敏感性，定量扩增编码酶的物种特异性mRNA 调节代谢流。为了设计必要的用于PCR的引物，编码酶，己糖激酶，磷酸果糖激酶、磷酸葡萄糖异构酶和柠檬酸盐合成酶将被克隆，利用在遗传上保守的这些酶的主要序列。物种特异性引物将从这些cDNA序列设计并用于定量地只从C. Finmarchicus确定环境参数对酶水平的影响表情 C.芬马尔基库斯出席在一个分类学上复杂的浮游动物组合中，通过同时测量特定浓度进行归一化生理上不变的基因，比如肌动蛋白。结果将相当于每重量C的酶。芬马尔基库斯队与更传统的生理学研究相比。