摘 要

研究几种食品加工方式对提取安康鱼肉DNA的影响。方法：以实时荧光PCR方法检测安康鱼成分标准为例，采取水煮，微波，油炸3种处理方式，选用试剂盒提取DNA，利用核酸蛋白测定仪测定其在260 nm,280 nm,230 nm处的吸光度，通过A260来计算核酸浓度，A260/A280和A260/A230来评估核酸的纯度。

以提取的DNA为模板，进行实时荧光PCR扩增。通过Ct值判断不同加工方式对实时荧光PCR结果的影响。结果：水煮，微波，油炸三种加工方式中，油炸对DNA的影响最明显，当在高温下油炸时间过久会导致鱼肉焦糖化反应及蛋白质变性严重，变成具有一定孔隙的焦状物，此时鱼肉在提取DNA的过程中难以消化，造成DNA提取量少，最终导致实时荧光PCR结果假阴性。而水煮，微波两种加工方式即使处理时间过久也没有对实时荧光PCR结果产生明显影响。

结论：研究表明，在鱼制品使用实时荧光PCR方法检测时，应充分考虑加工方式对鱼制品DNA的影响。

关键词：食品加工 安康鱼 DNA提取 实时荧光PCR

Effect of several food processing methods on DNA quality of Lophius lituion

Abstract

Objective To study the impact of several common food processing methods on DNA of Lophius lituion. Methods In this study, we used the standard of real-time PCR method for detecting DNA quality of Lophius lituion as an example. Lophius lituion was cooked at three cooking ways(water-boiling, microwave, frying). DNA was extracted by commercial DNA extraction kits. The purity of the DNA obtained from the samples in this study was calculated according to A260/A280 and A260/A230 values.

The concentration of DNA can be calculated by A260. The extracted DNA was amplification by real-time PCR. The influence of different processing methods on real-time PCR results was judged by Ct value. Results Frying was the most obvious effect on DNA in three kinds of processing methods. If the frying time at high temperature for too long, it will lead to the occurrence of serious caramelization and protein denaturation of Lophius lituion. Fish was turned into a porous coke. At this point, the fish in the process of extracting DNA is difficult to digest, what is resulting in less extraction of DNA and eventually lead to false negative results in real-time PCR. However, boiling, microwave processing has no significant impact on the results of real-time PCR, even if the processing time is too long.

Conclusion The study showed that the effect of processing methods on DNA of fish products should be fully considered when using real time PCR method for detecting fish products.

KEY WORDS: food processing; Lophius lituion; DNA extraction; Real-time PCR

目 录

摘 要 I

Abstract II

第一章 文献综述 4

1.1选题背景 4

1.2研究现状及分析 4

1.2.1SDS 2

1.2.2 CTAB 2

1.2.3试剂盒法 2

1.2.4酚一氯仿法 3

1.3研究目的及意义 3

第二章 实验材料与方法 5

2.1实验材料 5

2.1.1样品 5

2.1.2 仪器与试剂 5

2.2实验方法 5

2.2.1 样品处理 5

2.2.2 DNA提取 5

2.2.3DNA浓度及纯度的测定 6

2.2.4 实时荧光PCR检测 6

第三章 结果与讨论 7

3.1 不同温度下煮制不同时间对安康鱼肉DNA的影响 7

3.2微波加热对安康鱼肉DNA的影响 9

3.3不同温度下油炸不同时间对安康鱼肉DNA的影响 10

3.3.1不同油温在食品工艺上的加工方式区别 10

3.3.2 130℃油炸不同时间对安康鱼肉DNA的影响 10

3.3.3 160℃油炸不同时间对安康鱼肉DNA的影响 11

3.3.4 190℃油炸不同时间对安康鱼肉DNA的影响 12

3.3.5不同温度油炸对安康鱼肉DNA浓度及实时荧光PCR结果的影响 13

第四章 结论与展望 15

4.1结论 15

4.2展望 15

参考文献 16

致 谢 19

第一章 文献综述

1.1选题背景

近年来我国水产加工业发展迅速。水产制品层出不穷，安康鱼作为高营养价值的一个鱼种,在日本的几百年前安康鱼就是很高级的贡品，有着“东部吃安康、西部吃河豚”的美名。安康鱼肉除了低热量、低脂肪，还富含维生素A、D、E等。其中每100克的鱼肉中含0.8克左右的脂肪，且热量只有87大卡；营养丰富且胆固醇含量低，有增强体质、提高身体免疫力、预防肝脏疾病等功效。其鱼肉中具有抗氧化的成分，所含的胶质更是现代女性美容护肤提高身体年轻态的补品。

安康鱼的销量近些年在中国的市场逐步增长，每年销售安康鱼的数量也在水产品中占据了的较大份额。安康鱼在中国市场中的销售除了传统意义上的鲜鱼出售还包括了：鱼丸、鱼片、鱼豆腐等鱼类制品的销售。对于加工到鱼糜程度的肉类制品，我们无法通过传统的形态学来鉴别的安康鱼制品，我们只能进行分子生物学鉴定，其中PCR技术应用广泛。