Nonhematopoietically Derived DNA Is Shorter than Hematopoietically Derived DNA in Plasma: A Transplantation Model

Abstract

BACKGROUND Plasma DNA is predominantly hematopoietic in origin. The size difference between maternal- and fetal-derived DNA in maternal plasma prompted us to investigate whether there was any discrepancy in molecular size between hematopoietically and nonhematopoietically derived DNA in plasma. METHODS Plasma DNA samples from 6 hematopoietic stem cell transplant recipients and 1 liver transplant recipient were analyzed by massively parallel paired-end sequencing. The size of each fragment was deduced from the alignment positions of the paired reads. In sex-mismatched transplant recipients, the reads from chromosome Y were used as markers for the male donor/recipient. For other transplant recipients, the reads of the donor- and recipient-specific alleles were identified from the single-nucleotide polymorphism genotypes. RESULTS In male patients receiving female hematopoietic stem cells, more chromosome Y–derived DNA molecules (nonhematopoietically derived) were ≤150 bp than the autosome-derived ones (mainly hematopoietically derived) (median difference, 9.9%). In other hematopoietic stem cell transplant recipients, more recipient-specific DNA molecules (nonhematopoietically derived) were ≤150 bp than the donor-specific ones (hematopoietically derived) (median difference, 14.8%). In the liver transplant recipient, more donor-derived DNA molecules (liver derived) were ≤150 bp than the recipient-derived ones (mainly hematopoietically derived) (difference, 13.4%). The nonhematopoietically derived DNA exhibited a reduction in a 166-bp peak compared with the hematopoietically derived DNA. A 10-bp periodicity in size distribution below approximately 143 bp was observed in both DNA populations. CONCLUSIONS Massively parallel sequencing is a powerful tool for studying posttransplantation chimerism. Plasma DNA molecules exhibit a distinct fragmentation pattern, with the nonhematopoietically derived molecules being shorter than the hematopoietically derived ones.

Noninvasive Prenatal Methylomic Analysis by Genomewide Bisulfite Sequencing of Maternal Plasma DNA

“…3 in the online Data Supplement). These data are consistent with our previous findings that hematopoietic cells are the predominant source of DNA in human plasma (23 ).…”

“…We used paired-end sequencing to determine the lengths of plasma DNA molecules (23,32 ). In a previous study, we showed that plasma DNA molecules had sizes that approximated those of mononucleosomes and that fetal DNA molecules were shorter than maternal ones (32 ).…”

Cell-Free DNA: An Upcoming Biomarker in Transplantation

“…Different mechanisms accounting for DNA release in the circulation have been postulated, including apoptosis (23)(24)(25)(26), necrosis (26), and active secretion (27)(28)(29). Observations in gender-mismatched bone marrow and solid organ transplantation models point to hematopoietic cells as primary source of cfDNA in the plasma, with nonhematopoietic cells accounting for a minority of the free circulating DNA (24,30,31).…”

“…Observations in gender-mismatched bone marrow and solid organ transplantation models point to hematopoietic cells as primary source of cfDNA in the plasma, with nonhematopoietic cells accounting for a minority of the free circulating DNA (24,30,31). After crossing the kidney barrier by glomerular filtration, low molecular weight DNA fragments can appear in the urine, called ''transrenal DNA'' (Tr-DNA), whereas higher molecular weight DNA is derived from cells shed from the urinary tract or other urinary cells (32,33) ( Figure 1 and Table 1).…”

Use of Copy Number Deletion Polymorphisms to Assess DNA Chimerism

“…PCR primers were designed to locate within (i.e., internal PCR) and flank the deleted region (i.e., external PCR) of each CND marker in the panel. Short amplicons, ranging in size from 58 to 74 bp (mean 65 bp) were chosen for the internal PCRs so that they would be suitable for quantification of plasma ccfDNA, which is highly fragmented (11 ).…”