Universal Probe System for DNA Fingerprinting

DNA 指纹识别通用探针系统

• 批准号: 04670358
• 负责人: TAMAKI Yoshihiro
• 金额: $ 1.34万
• 依托单位: Oita Medical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04670358/
• 关键词: DNA fingerprint; Universal probe; Paternity testing

DNA fingerprinting requires the use of a specific DNA probe. A universal probe system is a combination of an unlabeled primary probe that has been inserted in a cloning vector and a labeled secondary probe that is specific to the vector. This system is time- and labor-saving in that one does not need to label probes every time one uses them, as long as they are inserted in the same vector.As first step to a universal probe system, we isolated three DNA fingerprinting probes from a subgenomic library that had been constructed by insertion into the phagemid pUC118 of 1-2 kb fragments from the EcoRI digest of DNA from human myeloma cells (named YA). Sequencing of one of the probes, termed YA1.8 after the length of the insert, revealed that although it contained no apparent repetitive sequence, it detected DNA fingerprints. This suggests that the probe is radically different from the known minisatellite probes ; therefore, we registered it with DDBJ under accession no.D29635.Next, we isolated single-stranded DNAs of recombinant phagemids, and hybridized them with southern blots of DNAs from father-mother-child trios or unrelated individuals. We were able to detect the DNA hybrids, that is, DNA fingerprints, with a commercially available, alkaline phosphatase-labeled secondary probe or a digoxigenin-labeled universal sequencing primer.

DNA指纹需要使用特定的DNA探针。通用探针系统是已插入克隆载体和特定于载体的标记的二次探针的未标记初级探针的组合。该系统是时间和延长的，因为只要将它们插入同一矢量，就不需要标记探针。作为通用探针系统的第一步，我们将三个DNA指纹探针从插入中插入的puc118的eCori Myer Myer Intra构成的亚基库中的三个DNA指纹探针（从1-2 KB的ecori群中构成）（是的）。插入长度后，其中一个探针的测序称为Ya1.8，表明尽管没有明显的重复序列，但它检测到DNA指纹。这表明探针与已知的微型片探针根本不同。因此，我们在登录号D29635下向DDBJ进行了注册，我们隔离了重组吞噬剂的单链DNA，并用父亲基尔德三个人或无关的个体与DNA的Southern DNA印迹杂交它们。我们能够检测到DNA杂种，即DNA指纹，并具有市售的碱性磷酸酶标记的二次探针或二高氧素标记的通用测序引物。

项目成果

Y.Tamaki: "Isolation of VNTR markers by anchored PCR of DNA fragments purified with immobilized oilignucleotide (in Japanese)." DNA Polymorphism. 1. 81-83 (1993)

Y.Tamaki：“通过固定化油寡核苷酸纯化的 DNA 片段的锚定 PCR 来分离 VNTR 标记（日语）。”

岸田 哲子: "部分的DNAライブラリーの選別によるVNTR型プローブの効率的単離" DNA多型研究の新しい展開. 1. 84-86 (1993)

Tetsuko Kishida：“通过筛选部分DNA文库有效分离VNTR型探针”DNA多态性研究的新进展1. 84-86 (1993)。

T.Kishida: "Efficient isolation of VNTR-probes from subgenomic library (in Japanese)." DNA Polymorphism. 1. 81-83 (1993)

T.Kishida：“从亚基因组文库中有效分离 VNTR 探针（日语）。”

玉置嘉広: "固相化オリゴヌクレオチドで精製したDNA断片のanchored PCRによるVNTRマーカーの単離" DNA多型研究の新しい展開. 1. 81-83 (1993)

Yoshihiro Tamaki：“通过固定寡核苷酸纯化的 DNA 片段的锚定 PCR 来分离 VNTR 标记”DNA 多态性研究的新进展 (1993)。

岸田哲子: "部分的DNAライブラリーの選別によるVNTR型プローブの効率的単離" DNA多型研究の新しい展開. 1. 84-86 (1993)

Tetsuko Kishida：“通过筛选部分DNA文库有效分离VNTR型探针”DNA多态性研究的新进展1. 84-86 (1993)。