Biomedical subjects

M N Gabriel

Publications and source records attributed to M N Gabriel.

3 recordsLinked to original sources

PubMed2001

Improved MtDNA sequence analysis of forensic remains using a "mini-primer set" amplification strategy.

Mitochondrial DNA (mtDNA) analysis of highly degraded skeletal remains is often used for forensic identification due largely to the high genome copy number per cell. Literature from the "ancient DNA" field has shown that highly degraded samples contain populations of intact DNA molecules that are severely restricted in size (1-4). Hand et al. have demonstrated the targeting and preferential amplification of authentic human DNA sequences with small amplicon products of 150 bp or less (1,2). Given this understanding of ancient DNA preservation and amplification, we report an improved approach to forensic mtDNA analysis of hypervariable regions 1 and 2 (HV1/HV2) in highly degraded specimens. This "mini-primer set" (MPS) amplification strategy consists of four overlapping products that span each of the HV regions and range from 126 to 170 bp, with an average size of 141 bp. For this study, 11 extracts representing a range of sample quality were prepared from nonprobative forensic specimens. We demonstrate a significant increase in MPS amplification success when compared to testing methods using approximately 250 bp amplicons. Further, 16 of 17 independent amplifications previously "unreported" due to mixed sequences provided potentially reportable sequence data from a single, authentic template with MPS testing.

Bone and Bones↗

PubMed2001

Population variation of human mitochondrial DNA hypervariable regions I and II in 105 Croatian individuals demonstrated by immobilized sequence-specific oligonucleotide probe analysis.

AIM: To detect sequence variation in 105 Croatian individuals by the use of duplex polymerase chain reaction amplification of full-length hypervariable region I and II (HVI/HVII) products and subsequent hybridization to a linear array of 27 immobilized sequence-specific oligonucleotide (SSO) probes, which targets six regions within HVI and HVII, and two additional sites, 189 and 16093. METHODS: Chelex-extracted bloodstains were used for amplification of HV regions. In all cases, a single robust amplification was sufficient for immobilized SSO probe typing and subsequent direct sequence analysis for both HVI and HVII. This method, suitable for a range of forensic samples (including shaft portions of single hairs), was also applied to the analysis of 18 skeletal elements recovered from a mass grave. Using a panel of immobilized SSO probes, we have developed a rapid screening approach to mitochondrial DNA (mtDNA) haplotyping before direct sequence analysis. RESULTS: We established a reference sequence database of mtDNA haplotypes for 105 randomly selected Croatian individuals. Fifty different mitotypes were observed (33 unique). The most frequent mitotypes occurred 18 times or approximately 17.1% [111111 189 (A) 16093 (T)] and 11 times or approximately 10.5% [131111 189 (A) 16093 (T)]; all other mitotypes occurred 5% or less. The corresponding genetic diversity value for this database was approximately 0.952. The usefulness of establishing an mtDNA reference database with immobilized SSO probe testing has been demonstrated by determining the strength of a match comparison obtained for one skeletal element and a corresponding maternal reference from 18 specimens recovered from a mass grave. CONCLUSION: The sequence variation detected by the panel of immobilized SSO probes is sufficiently diverse to be used for identification of human skeletal remains from mass graves. The immobilized SSO typing strip targets polymorphic regions within HVI and HVII and is a useful identification tool for mass grave and mass disaster analysis, as well as for criminal casework testing.

Base Sequence↗

PubMed1997

Repression of human immunodeficiency virus type 1 through the novel cooperation of human factors YY1 and LSF.

A subpopulation of stably infected CD4+ cells capable of producing virus upon stimulation has been identified in human immunodeficiency virus (HIV)-positive individuals (T.-W. Chun, D. Finzi, J. Margolick, K. Chadwick, D. Schwartz, and R. F. Siliciano, Nat. Med. 1:1284-1290, 1995). Few host factors that directly limit HIV-1 transcription and could support this state of nonproductive HIV-1 infection have been described. YY1, a widely distributed human transcription factor, is known to inhibit HIV-1 long terminal repeat (LTR) transcription and virus production. LSF (also known as LBP-1, UBP, and CP-2) has been shown to repress LTR transcription in vitro, but transient expression of LSF has no effect on LTR activity in vivo. We report that both YY1 and LSF participate in the formation of a complex that recognizes the initiation region of the HIV-1 LTR. Further, we have found that these factors cooperate in the repression of LTR expression and viral replication. This cooperative function may account for the divergent effects of LSF previously observed in vitro and in vivo. Thus, the cooperation of two general cellular transcription factors may allow for the selective downregulation of HIV transcription. Through this mechanism of gene regulation, YY1 and LSF could contribute to the establishment and maintenance of a population of cells stably but nonproductively infected with HIV-1.

Cell Line↗