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

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。

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.