Comparison of short tandem repeat and variable number tandem repeat genetic markers for quantitative determination of allogeneic bone marrow transplant engraftment

Abstract

Summary:Variable number tandem repeats (VNTRs) were among the first genetic markers used to quantitate bone marrow transplant engraftment. STRs, which are standard tools for genotyping in parentage testing 6 and forensic human identity testing, 7 have recently been used for the quantitative analysis of BMT engraftment. [8][9][10][11] STRs provide an excellent tool for this purpose because of their high degree of polymorphism and relatively short length. Furthermore, STR-based human identity kits are commercially available, which eliminates the need for developing the methodology in each individual laboratory. Several of the commercial STR human identity kits have been optimized for automated DNA sequencers which enables data collection and analysis by sophisticated software. Commercial kits for VNTR analysis currently are not available. Although STRs are now widely used in BMT testing, only one direct comparison of VNTR and STR engraftment results has been published. 11 The purpose of this study is to validate STRs in BMT engraftment analysis by comparing VNTR and STR results for the same sets of post-transplant samples from patients who had undergone allogeneic BMT. Sixteen post-transplant samples from four allogeneic BMT patients are included in this correlation study. Patient samples previously analyzed by the VNTR method at the FairviewUniversity Medical Center Molecular Diagnostics Laboratory were chosen to cover the full range of engraftment from near 0% to 100%. STR analysis was then performed on the same samples in a blinded fashion at the Central Arkansas Veterans Healthcare System Molecular Diagnostics Laboratory. Good quantitative correlation was found between STR and VNTR results in samples from all four patients. STR markers were informative in one patient for whom PCR-based VNTR markers were not available. This study demonstrates that STR-based human identity testing kits are well suited for engraftment analysis.

Transplantation of ex-vivo culture-expanded parental haploidentical mesenchymal stem cells to promote engraftment in pediatric recipients of unrelated donor umbilical cord blood: results of a phase I–II clinical trial

“…The method of analysis was quantitative PCR of informative polymorphic variable number tandem repeat or short tandem repeat regions in the recipient and donor. 16,17 Results…”

Bone Marrow Transplant Engraftment Analysis with Loss of an Informative Allele

“…Stutter peaks of tetranucleotide STRs are 4 bp shorter than the main peak and usually have a peak area close to 5% of the main peak area. 1 In Figure 2A, stutter peaks amplified at the donor D3S1358 locus are present in the (13) position, derived from the D3S1358 (14) allele, and in the (16) position, derived from the D3S1358 (17) allele. Small stutter peaks amplified at the donor vWA locus are present in the (14) position, derived from the vWA (15) allele, and the (16) position, derived from the vWA (17) allele.…”

“…Fragment size and peak area data were determined by GeneScan and GenoTyper software (Applied Biosystems). 1 The D3S1358 and vWA loci show informative alleles that distinguish donor ( Figure 2A) from recipient ( Figure 2B). At the D3S1358 locus, donor and recipient are both heterozygous with no shared alleles.…”

“…1 Ratios of the peak areas of donor and recipient alleles at an informative locus are used to calculate the DNA percentages in post-transplant samples. Specific formulas for calculating the percentage of donor or recipient DNA are based on whether the informative loci are heterozygous or homozygous and whether the donor and recipient share any alleles.…”

“…1 After amplification, samples were analyzed on an ABI 373A DNA Sequencer (Applied Biosystems). Fragment size and peak area data were determined by GeneScan and GenoTyper software (Applied Biosystems).…”