Saha A, Husain S, Bamezai R
Human Genetics Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi, India.
DNA Cell Biol. 2000 Apr;19(4):219-26. doi: 10.1089/104454900314483.
Sequence analysis was carried out of a human clone pBA0.6 generated after exonuclease III/S1 nuclease digestion and subcloning of pCMM86 (GDB: 168382, D17S74), which was not available in the database. It revealed the presence of a reiterating core motif of 24mer GTGGGTGTGTTGGAGGGGGTGAGG, present 23 times, which was GC-rich and minisatellitic in nature. Genomic blots of HaeIII-digested human DNA, when hybridized with pBA0.6, generated a ladder of bands between 29.0 kb and 2.1 kb. Hybridization analyses of 88 unrelated individuals belonging to four regions of India using this probe revealed polymorphic bands which were individual specific. The probability of identity ranged from 5.07x10(-14) in Punjabis to 2.64x10(-16) in Bengalis and was found to be 3.06x10(-16) in UPites, whereas in the case of South Indians, it was 3.9x10(-15). Three sets of isomorphic bands at 29.0 kb, 2.4 kb, and 2.1 kb were common between the individuals of all the regions and served as internal markers. The 29.0-kb band was observed to be Homo sapiens specific. Construction of dendrograms based on the UPGMA method with Jaccard's coefficient values suggested less genetic similarity/high genetic diversity in all the population groups, indicating that the samples taken were random. Maximum likelihood estimates through the bootstrap sampling method showed that Punjabis, Bengalis, and UPites formed one cluster, whereas South Indians formed a separate cluster, altogether thus showing the proximity of these three population groups compared with that from South India. A preliminary study by Northern hybridization with pBA0.6 resulted in two transcripts of 0.63 kb and 0.29 kb. This finding was corroborated with RT-PCR results where 2 amplicons, matching the expected size of two open reading frames within the minisatellite sequence, were obtained. The role of the two transcripts from the minisatellite sequence is not clear as yet, and it is probable that these messages may not get translated because of the absence of a eukaryotic Kozak sequence around the initiator methionine in the pBA0.6 sequence.
对通过核酸外切酶III/S1核酸酶消化并亚克隆pCMM86（GDB：168382，D17S74）后产生的人类克隆pBA0.6进行了序列分析，该克隆在数据库中不可用。分析揭示了一个24聚体GTGGGTGTGTTGGAGGGGGTGAGG的重复核心基序，共出现23次，其本质上富含GC且为小卫星序列。用HaeIII消化的人类DNA进行基因组印迹，当与pBA0.6杂交时，产生了29.0 kb至2.1 kb之间的一系列条带。使用该探针对来自印度四个地区的88名无关个体进行杂交分析，发现了个体特异性的多态性条带。个体识别概率在旁遮普人中为5.07×10（-14），在孟加拉人中为2.64×10（-16），在北方邦人中为3.06×10（-16），而在南印度人中为3.9×10（-15）。29.0 kb、2.4 kb和2.1 kb的三组同形条带在所有地区的个体之间是共有的，并作为内部标记。观察到29.0 kb的条带是智人特有的。基于UPGMA方法和杰卡德系数值构建的树状图表明，所有种群组中的遗传相似性较低/遗传多样性较高，表明所采集的样本是随机的。通过自展抽样方法进行的最大似然估计表明，旁遮普人、孟加拉人和北方邦人形成一个聚类，而南印度人形成一个单独的聚类，因此总体上显示出这三个种群组与来自南印度的种群组相比更为接近。用pBA0.6进行Northern杂交的初步研究产生了0.63 kb和0.29 kb的两个转录本。这一发现得到了RT-PCR结果的证实，其中获得了2个扩增子，与小卫星序列内两个开放阅读框的预期大小相符。来自小卫星序列的这两个转录本的作用尚不清楚，并且由于pBA0.6序列中起始甲硫氨酸周围缺乏真核生物Kozak序列，这些信息可能无法翻译。
