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CN106905399B - 一种双链dna b-z构象相互转换的光控方法 - Google Patents

一种双链dna b-z构象相互转换的光控方法 Download PDF

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CN106905399B
CN106905399B CN201710189622.7A CN201710189622A CN106905399B CN 106905399 B CN106905399 B CN 106905399B CN 201710189622 A CN201710189622 A CN 201710189622A CN 106905399 B CN106905399 B CN 106905399B
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dna
conformation
double
stranded dna
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CN106905399A (zh
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刘奕侬
杜宇昊
周翔
何志勇
饶爽
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Wuhan University WHU
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    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H21/00—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
    • C07H21/04—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
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Abstract

本发明提供一种双链DNA B‑Z构象相互转换的光控方法，属于分子生物学和核酸化学领域。该方法首先通过DNA标准固相合成法，合成带有修饰鸟嘌呤短链DNA，然后通过365nm、254nm波长的光照，可逆的控制B型DNA和Z型DNA的相互转化。该方法利用光照可逆的控制DNA构象转化，应用中具有时空可控性。

Description

一种双链DNA B-Z构象相互转换的光控方法
技术领域
本发明属于分子生物学和核酸化学领域，涉及一种B型DNA和Z型DNA的相互转化的光控可逆控制的方法。
背景技术
DNA有三种主要构象：A-DNA、B-DNA和Z-DNA。B-DNA即为Watson和Click提出的右手螺旋模型，是大部分双螺旋DNA在生理条件下的构像。A-DNA也为右手螺旋，但是螺体较宽而短，大沟区较窄而深，小沟区较浅而宽。A型DNA可由B-DNA部分脱水得到，通常在非生理条件下形成，而在体内，A-DNA可能产生于DNA-RNA的杂交或DNA与酶的复合物。Z-DNA与上两种不同，是左手螺旋，磷酸基分布呈Z字型，只有一条大沟而无小沟。嘌呤-嘧啶交替排列的序列特别是d(GC)交替序列容易形成Z-DNA，如果其中C甲基化，则更易形成Z-DNA。Z型DNA可以被特定的蛋白所识别，而这一过程对转录的调控具有重要意义。B-DNA和Z-DNA已经在体内被直接观测。。
DNA的各种二级结构之间可以相互转变，对于生物大分子来说，这种转变具有重要生理功能。但是科学家们始终没有对Z-DNA的结构与其功能形成系统的认识，目前研究Z-DNA的手段都是通过Z-DNA结合蛋白去探究某些Z-DNA的功能，因此，本发明开发的光控可逆的控制B型DNA和Z型DNA的相互转化的方法，可以实现对生物大分子的操作和调控，对于相关生理作用的分子机制的研究显得尤为重要。
发明内容
本发明所要解决的技术问题在于提供一种具有时空可控性，并且对活体伤害小，基于修饰的鸟嘌呤利用光照控制双链DNA B-Z构象相互转化的方法。
本发明的技术方案是：
1)、利用DNA固相合成法，合成含有一个如通式(I)的修饰鸟嘌呤的短链DNA，然后与互补链退火后形成双链DNA；其中，R1为苯基、2-萘基或2-芴基中的一种；
2)在0.2-4摩尔/升的NaCl溶液中，用波长365nm光照10分钟，然后37度温浴6小时，DNA构象由B转向Z；用波长254nm光照Z构象的DNA10分钟，然后37度温浴6小时，可以实现DNA构象由Z转向B的可逆控制。
本发明的原理图见图1所示。
本发明的优点和有益效果在于：
1、本发明中对DNA构象的B-Z可逆转化的控制用到的是光照，具有时空可控性，并且对活体伤害小，利于原位控制。
2、本发明中的DNA构象的B-Z转化是可逆的，使得该方法非常适于研究B-Z转化相关生理功能的分子机制。
附图说明
图1DNA构型B-Z转化光控可逆控制的工作原理图
图2双链DNA不同NaCl浓度下CD光谱
图3 B-Z构型光控可逆转化
具体实施方式
通过以下详细说明结合附图可以进一步理解本发明的特点和优点。所提供的实施例仅是对本发明方法的说明，而不以任何方式限制本发明揭示的其余内容。
实施例1
1、本发明含有修饰鸟嘌呤的DNA合成
应用DNA固相合成法合成含有修饰鸟嘌呤的DNA(Angew.Chem.Int.Ed.2008,47,8839–8842)，其序列为CGCXCGCGCGC，X表示图1中结构修饰的鸟嘌呤，其互补DNA为普通序列GCGCGCGCGCG。在pH 7.0，5毫摩尔/升的磷酸钠缓冲溶液中退火，形成双链DNA。
2、NaCl的浓度对B-DNA到Z-DNA结构转变的影响
在10℃，DNA双链浓度为10μM的条件下，向溶液中加入NaCl，至终浓度分别为1，2，3，4，5摩尔/升，并分别测量溶液的CD谱图，结果如图2所示，可以看到295nm处的吸收值由正向负值转变，表明随着NaCl浓度增加，DNA构象由B向Z转化。
3、不同波长的光照控制DNA B-Z构型可逆转化
在双链DNA溶液中，如图3所示，加入NaCl至终浓度为3.5摩尔/升，测量CD谱图，为图3绿线；然后365nm光照10分钟，接着37度温浴6小时，测量CD值，为图3红线所示。然后波长254nm光照10分钟，然后37度温浴6小时，测量CD值，为图3浅蓝线所示。再365nm光照10分钟，接着37度温浴6小时，测量CD值，为图3深蓝线所示。然后波长254nm光照10分钟，然后37度温浴6小时，测量CD值，为图3粉红线所示。CD谱图的295nm处的吸收值随光照发生的改变，表明不同波长光照对DNA构象B-Z转化的可逆调控。

Claims (1)

1.一种基于修饰的鸟嘌呤利用光照控制双链DNAB-Z构象相互转化的方法，其特征在于，包括如下步骤：
1)利用DNA固相合成法，合成含有一个如(I)的修饰鸟嘌呤的短链DNA，然后与互补链退火后形成双链DNA；
2)在3.5摩尔/升的NaCl溶液中，用波长365nm光照10分钟，然后37度温浴6小时，DNA构象由B转向Z；用波长254nm光照Z构象的DNA10分钟，然后37度温浴6小时，可以实现DNA构象由Z转向B的可逆控制；
CN201710189622.7A 2017-03-27 2017-03-27 一种双链dna b-z构象相互转换的光控方法 Active CN106905399B (zh)

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Publication number Priority date Publication date Assignee Title
CN1212021A (zh) * 1996-01-23 1999-03-24 拉普吉恩公司 测定核酸分子序列的方法和组合物
WO2007107775A2 (en) * 2006-03-23 2007-09-27 The Queen's University Of Belfast Preparation of oligonucleotides with photoswitchable properties
CN102171368A (zh) * 2008-10-01 2011-08-31 皇家飞利浦电子股份有限公司 在载体上固定化核酸的方法

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1212021A (zh) * 1996-01-23 1999-03-24 拉普吉恩公司 测定核酸分子序列的方法和组合物
WO2007107775A2 (en) * 2006-03-23 2007-09-27 The Queen's University Of Belfast Preparation of oligonucleotides with photoswitchable properties
CN102171368A (zh) * 2008-10-01 2011-08-31 皇家飞利浦电子股份有限公司 在载体上固定化核酸的方法

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