Su Qiang, Nöll Gilbert
Nöll Junior Research Group, Organic Chemistry, Chem. Biol. Dept., Faculty IV, Siegen University, Adolf-Reichwein-Str. 2, 57068, Siegen, Germany.
Chemistry. 2017 Jan 12;23(3):696-702. doi: 10.1002/chem.201604652. Epub 2016 Nov 30.
The orientation of surface-grafted dsDNA assemblies relative to the surface depends strongly on the nature of the employed thiol anchor. This was shown by ssDNA capture probe strands of 20 bases grafted to a gold surface by three dithiane rings or a single mercaptohexyl group. The capture probe strands were hybridized to one end of complementary ssDNA strands (target) comprising 40, 60, or 80 bases (T , T , and T ). At the other end of the targets a fluorophore-labeled reporter probe ssDNA strand of 20 bases was hybridized. To stiffen the DNA assemblies, the targets T and T were further hybridized to ssDNA patches of 20 or 40 bases. Whether the fluorescence intensity, and thus the distance between surface and fluorophore, increases or decreases with increasing target length depends on the thiol anchor. Attempts were made to heal the nicks that are present in the formed dsDNA assemblies by ligation. For enzymatic ligation, the presence of a phosphate at the 5'-end of the reporter probe and a patch is required, which may also influence the fluorescence intensity.
表面接枝的双链DNA组装体相对于表面的取向在很大程度上取决于所采用的硫醇锚定基团的性质。这一点通过20个碱基的单链DNA捕获探针链得到了证明，这些探针链通过三个二硫杂环或单个巯基己基基团接枝到金表面。捕获探针链与包含40、60或80个碱基（T₄₀、T₆₀和T₈₀）的互补单链DNA链（靶标）的一端杂交。在靶标的另一端，杂交了一条20个碱基的荧光团标记的报告探针单链DNA。为了使DNA组装体变硬，靶标T₆₀和T₈₀进一步与20或40个碱基的单链DNA片段杂交。荧光强度以及表面与荧光团之间的距离是否随靶标长度增加而增加或减少取决于硫醇锚定基团。人们试图通过连接来修复形成的双链DNA组装体中存在的切口。对于酶促连接，报告探针和片段的5'端需要有磷酸基团，这也可能影响荧光强度。
