A 13-plex of tetra- and penta-STRs to identify zebrafish.
A 13-plex of tetra- and penta-STRs to identify zebrafish.
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用于识别斑马鱼的 13 重四重和五重 STR。
DOI：
10.1038/s41598-020-60842-5
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发表时间：
2020
影响因子：
4.6
通讯作者：
Cibelli,JoseB
中科院分区：
文献类型：
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作者：
Venta,PatrickJ;Nguyen,AnthonyK;Senut,Marie-Claude;Poulos,WilliamG;Prukudom,Sukumal;Cibelli,JoseB
斑马鱼（Danio rerio）已成为生物医学研究中主要的脊椎动物模式生物之一。然而，该模式的某些方面需要改进。其中之一是能够通过DNA分析来识别个体鱼和鱼线。虽然许多二核苷酸短串联重复序列（diSTR）标记可用于此目的和类似的目的，但它们具有某些缺点，如过度的聚合酶滑移（“口吃”），这导致自动基因分型和跨实验室比较的困难。在这里，我们报告了一个13重的四核苷酸和五核苷酸STR（tetraSTR和pentaSTR，分别），具有低口吃的发展。该系统使用廉价的通用引物标记系统，如果需要，可以很容易地转换为直接标记系统。在三个斑马鱼品系（NHGRI-1、kca 33 Tg和kca 66 Tg，最初从ZIRC获得）中检查了该13重。这些高度近交系的平均观察杂合性（Ho）和预期杂合性（He）分别为0.291和0.359，这与diSTR的结果非常相似。根据Genalex软件包计算，所有测试鱼的同一性概率（PI）为2.1 × 10− 5，同胞鱼的PI（PIsib）为6.4 × 10−3。百分之九十的鱼被正确地识别出各自的菌株。还证明了该组可用于确认双单倍体细胞系。这种多重应该找到多种用途，以提高使用斑马鱼模型的研究的准确性和再现性。
The zebrafish speciesDanio reriohas become one of the major vertebrate model organisms used in biomedical research. However, there are aspects of the model that need to be improved. One of these is the ability to identify individual fish and fish lines by DNA profiling. Although many dinucleotide short tandem repeat (diSTR) markers are available for this and similar purposes, they have certain disadvantages such as an excessive polymerase slippage (“stutter”) that causes difficulties in automated genotyping and cross-laboratory comparisons. Here we report on the development of a 13-plex of tetranucleotide and pentanucleotide STRs (tetraSTRs and pentaSTRs, respectively) that have low stutter. The system uses an inexpensive universal primer labelling system, which can easily be converted to a direct labeling system if desired. This 13-plex was examined in three zebrafish lines (NHGRI-1, kca33Tg, and kca66Tg, originally obtained from ZIRC). The average observed heterozygosity (Ho) and expected heterozygosity (He) in these highly inbred lines were 0.291 and 0.359, respectively, which is very similar to what has been found with diSTRs. The probability of identity (PI) for all fish tested was 2.1 × 10−5and the PI for siblings (PIsib) was 6.4 × 10−3, as calculated by the Genalex package. Ninety percent of the fish tested were correctly identified with their respective strains. It is also demonstrated that this panel can be used to confirm doubled-haploid cell lines. This multiplex should find multiple uses for improving the accuracy and reproducibility of studies using the zebrafish model.