Kao P C, Beyer C F, Lang C A
Biochem J. 1976 Feb 15;154(2):471-80. doi: 10.1042/bj1540471.
In previous experiments on growth and aging in the yellow-fever mosquito, Aedes aegypti, a low mol. wt. (500000) DNA species was found in the supernatant fraction after ultracentrifugation of homogenates of rapidly-growing larvae. This DNA species, "sDNA", constituted 30-40% of total DNA in 2-4-day-old larvae, but was less than 5% in older larvae, pupae and adults. We have now isolated and characterized sDNA and initiated experiments to determine its metabolic role. Isolated sDNA has the same physical and chemical characteristics as bulk DNA, "pDNA", and differs only in size. In CsCl isopycnic centrifugation the buoyant densities of sDNA and pDNA were 1.700 and 1.697 g/cm3 respectively. The "melting" temperature of both DNA species was 84 degrees C. Base compositions calculated from these data and other methods were 38.9 mol% of guanine-plus-cytosine for sDNA, and 38.5 for pDNA. Also, the size of newly-synthesized DNA was investigated by pulse-labelling and pulse-chase experiments. In neutral sucrose gradients the labelled DNA component after a 2h pulse had a sedimentation coefficient of about 8S, but after a 4h pulse sedimented in a broad band from 10-19S. In alkaline sucrose gradients a single peak around 7S was observed for pulse times up to 4h. After a 6h pulse and a 1 day "chase", labelled DNA species had sedimentation coefficients ranging from 10-15S in alkaline sucrose, and after a 2-day chase the values were 17-31S, similar to those of pDNA under alkaline conditions. These results suggest that sDNA represents an intermediate form in the replication of DNA in mosquito larvae.
在先前关于黄热病蚊子埃及伊蚊生长和衰老的实验中，在快速生长幼虫匀浆超速离心后的上清液部分发现了一种低分子量（500000）的DNA物种。这种DNA物种，即“sDNA”，在2至4日龄幼虫的总DNA中占30 - 40%，但在老龄幼虫、蛹和成虫中占比不到5%。我们现已分离并鉴定了sDNA，并启动了实验以确定其代谢作用。分离出的sDNA具有与大量DNA（“pDNA”）相同的物理和化学特性，仅在大小上有所不同。在CsCl等密度离心中，sDNA和pDNA的浮力密度分别为1.700和1.697 g/cm³。两种DNA物种的“熔解”温度均为84℃。根据这些数据和其他方法计算得出的碱基组成，sDNA中鸟嘌呤加胞嘧啶为38.9摩尔%，pDNA为38.5摩尔%。此外，通过脉冲标记和脉冲追踪实验研究了新合成DNA的大小。在中性蔗糖梯度中，2小时脉冲后标记的DNA组分沉降系数约为8S，但4小时脉冲后在10 - 19S的宽条带中沉降。在碱性蔗糖梯度中，脉冲时间长达4小时时观察到一个约7S的单峰。6小时脉冲和1天“追踪”后，标记的DNA物种在碱性蔗糖中的沉降系数范围为10 - 15S，2天追踪后值为17 - 31S，与碱性条件下pDNA的值相似。这些结果表明，sDNA代表了蚊子幼虫DNA复制中的一种中间形式。
