Sampson S L, Warren R M, Richardson M, van der Spuy G D, van Helden P D
Department of Medical Biochemistry, University of Stellenbosch Medical School, Tygerberg.
Tuber Lung Dis. 1999;79(6):349-59. doi: 10.1054/tuld.1999.0218.
The insertion sequence IS6110 is widely used as a DNA fingerprinting probe for the classification of Mycobacterium tuberculosis strains. This study has focused on the characterization of regions disrupted by insertion of the IS6110 element.
To characterize IS6110 insertion loci in clinical isolates of M. tuberculosis, in terms of their genomic location and genetic identity, to ascertain whether IS6110 transposition could be a mechanism driving phenotypic change.
Thirty-three IS6110 insertion loci were cloned from 8 clinical isolates of M. tuberculosis. Clones representing DR locus insertions were identified by hybridization (n = 4), and all other clones were characterized by DNA sequencing (n = 29). The sequence data was analyzed in conjunction with that of 43 other insertion loci identified in published literature and DNA sequence databases.
The 76 sequences analyzed represented 66 unique insertion loci (including 9 unique insertions into the ipl locus). When mapped to the H37Rv genome, the majority of unique insertion loci demonstrated disruption of coding regions by IS6110 (n = 42; including the ipl insertions), while the remainder either occurred within intergenic regions (n = 17), or could not be mapped to the H37Rv genome sequence (n = 7). Mapping of the insertion loci reveals distribution throughout the chromosome, with isolated preferential insertion loci.
This study has demonstrated the occurrence of 66 unique IS6110 insertion loci dispersed throughout the M. tuberculosis genome, with an unexpectedly high incidence of IS6110 insertions occurring within coding regions. However, the IS6110-mediated coding region disruptions identified here may only have limited impact on phenotype, as most of the coding regions disrupted are members of multiple gene families. Disruption of individual members of a family of genes may have no effect on phenotype or could have a minor or major impact, depending on the specificity and activity of the encoded protein.
插入序列IS6110被广泛用作结核分枝杆菌菌株分类的DNA指纹图谱探针。本研究聚焦于被IS6110元件插入所破坏区域的特征分析。
从基因组位置和遗传同一性方面对结核分枝杆菌临床分离株中的IS6110插入位点进行特征分析，以确定IS6110转座是否可能是驱动表型变化的一种机制。
从8株结核分枝杆菌临床分离株中克隆出33个IS6110插入位点。通过杂交鉴定代表DR位点插入的克隆（n = 4），所有其他克隆通过DNA测序进行特征分析（n = 29）。结合已发表文献和DNA序列数据库中鉴定的其他43个插入位点的序列数据进行分析。
分析的76个序列代表66个独特的插入位点（包括9个插入到ipl位点的独特插入）。当定位到H37Rv基因组时，大多数独特的插入位点显示编码区被IS6110破坏（n = 42；包括ipl插入），其余的要么发生在基因间区域（n = 17），要么无法定位到H37Rv基因组序列（n = 7）。插入位点的定位揭示了其在整个染色体上的分布，存在孤立的优先插入位点。
本研究证明了66个独特的IS6110插入位点分散在结核分枝杆菌基因组中，且在编码区内发生IS6110插入的发生率出乎意料地高。然而，此处鉴定的IS6110介导的编码区破坏可能仅对表型有有限影响，因为大多数被破坏的编码区是多个基因家族的成员。基因家族中单个成员的破坏可能对表型没有影响，也可能有轻微或重大影响，这取决于所编码蛋白质的特异性和活性。
