A Consensus Polymerase Chain Reaction–Oligonucleotide Hybridization Approach for the Detection of Chromosomal Translocations in Pediatric Bone and Soft Tissue Sarcomas

Abstract Ewing sarcoma/primitive neuroectodermal tumor is classically a tumor of the soft tissue or bone in children and young adults, but several cases have been described in patients of all ages. Within the last decade, the clinicopathologic spectrum of Ewing sarcoma/primitive neuroectodermal tumor has been markedly expanded by recognition that the tumor may also have a visceral origin. We describe a case of primitive neuroectodermal tumor arising in the stomach of a 66-year-old woman. The neoplasm was excised using a radical surgical procedure. Microscopically, the tumor was made up of solid nests and sheets of round cells. Immunohistochemically, the tumor cells showed immunoreactivity for CD99, S100, neuron-specific enolase, and vimentin. A multiplex real-time polymerase chain reaction assay detected an EWS-ERG fusion. To our knowledge, this is the first description of a primitive neuroectodermal tumor arising in the stomach.

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16S rRNA-targeted polymerase chain reaction and oligonucleotide hybridization to screen for Azoarcus spp., grass-associated diazotrophs.

Applied and Environmental Microbiology ◽

10.1128/aem.59.11.3816-3824.1993 ◽

1993 ◽

Vol 59 (11) ◽

pp. 3816-3824 ◽

Cited By ~ 31

Author(s):

T Hurek ◽

S Burggraf ◽

C R Woese ◽

B Reinhold-Hurek

Keyword(s):

Polymerase Chain Reaction ◽

16S Rrna ◽

Chain Reaction ◽

Oligonucleotide Hybridization ◽

Polymerase Chain

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Detection ofRAS mutations in archival testicular germ cell tumors by polymerase chain reaction and oligonucleotide hybridization

Genes Chromosomes and Cancer ◽

10.1002/gcc.2870050204 ◽

1992 ◽

Vol 5 (2) ◽

pp. 109-118 ◽

Cited By ~ 32

Author(s):

Judd W. Moul ◽

Sheila M. Theune ◽

Esther H. Chang

Keyword(s):

Polymerase Chain Reaction ◽

Germ Cell ◽

Germ Cell Tumors ◽

Testicular Germ Cell Tumors ◽

Chain Reaction ◽

Testicular Germ Cell ◽

Oligonucleotide Hybridization ◽

Polymerase Chain

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Diagnotic Significance of Detection of Fusion Genes by Reverse Transcriptase-Polymerase Chain Reaction in Soft Tissue Tumors

Orthopedics & Traumatology ◽

10.5035/nishiseisai.58.473 ◽

2009 ◽

Vol 58 (3) ◽

pp. 473-477

Author(s):

Kensaku Yamaga ◽

Hideki Yamashita ◽

Koji Endo ◽

Mitsuhiko Osaki ◽

Takeshi Minamizaki ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Soft Tissue ◽

Reverse Transcriptase ◽

Soft Tissue Tumors ◽

Fusion Genes ◽

Chain Reaction ◽

Polymerase Chain

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Amplification by the polymerase chain reaction of hypervariable regions of the human genome for evaluation of chimerism after bone marrow transplantation

Blood ◽

10.1182/blood.v77.7.1607.bloodjournal7771607 ◽

1991 ◽

Vol 77 (7) ◽

pp. 1607-1615 ◽

Cited By ~ 1

Author(s):

L Ugozzoli ◽

P Yam ◽

LD Petz ◽

GB Ferrara ◽

RE Champlin ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Marrow Transplantation ◽

Tandem Repeats ◽

Limit Of Detection ◽

Minor Component ◽

Restriction Enzymes ◽

Mixed Chimerism ◽

Chain Reaction ◽

Oligonucleotide Hybridization ◽

Polymerase Chain

We combined the polymerase chain reaction (PCR) with oligonucleotide hybridization as a novel and sensitive technique to evaluate posttransplant chimerism. Specific oligonucleotides for hybridization were synthesized homologous to tandemly repetitive core sequences of regions with a variable number of tandem repeats (VNTRs). Polymorphisms at such loci result from allelic differences in the number of repeats. Primers flanking the repeat region of each of the corresponding VNTRs were used for amplification. Recipient and donor pretransplant DNA and recipient posttransplant DNA were amplified. The resultant fragments were analyzed after gel electrophoresis either by hybridization in-gel or after Southern transfer. To confirm our findings, we also performed standard assays of restriction fragment length polymorphisms (RFLPs). Evaluation of 13 selected cases indicated mixed chimerism (4), complete chimerism (5), recurrence of leukemia (2), and endogenous repopulation of hematopoiesis (2) after marrow transplantation. Sensitivity of the method was determined by mixing various proportions of recipient and donor DNA; the limit of detection of the minor component in a mixture was 0.1%. PCR data correlated with RFLP data in all cases except two in which PCR proved more sensitive than RFLP. PCR amplification of VNTRs combined with oligonucleotide hybridization is a novel technique for documenting posttransplant chimerism and has advantages over RFLP analysis: high sensitivity, use of small amounts of DNA (250 ng), ease of preparation of DNA, elimination of need for restriction enzymes, and the ability to complete studies in 2 days.

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Amplification by the polymerase chain reaction of hypervariable regions of the human genome for evaluation of chimerism after bone marrow transplantation

Blood ◽

10.1182/blood.v77.7.1607.1607 ◽

1991 ◽

Vol 77 (7) ◽

pp. 1607-1615 ◽

Cited By ~ 107

Author(s):

L Ugozzoli ◽

P Yam ◽

LD Petz ◽

GB Ferrara ◽

RE Champlin ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Marrow Transplantation ◽

Tandem Repeats ◽

Limit Of Detection ◽

Minor Component ◽

Restriction Enzymes ◽

Mixed Chimerism ◽

Chain Reaction ◽

Oligonucleotide Hybridization ◽

Polymerase Chain

Abstract We combined the polymerase chain reaction (PCR) with oligonucleotide hybridization as a novel and sensitive technique to evaluate posttransplant chimerism. Specific oligonucleotides for hybridization were synthesized homologous to tandemly repetitive core sequences of regions with a variable number of tandem repeats (VNTRs). Polymorphisms at such loci result from allelic differences in the number of repeats. Primers flanking the repeat region of each of the corresponding VNTRs were used for amplification. Recipient and donor pretransplant DNA and recipient posttransplant DNA were amplified. The resultant fragments were analyzed after gel electrophoresis either by hybridization in-gel or after Southern transfer. To confirm our findings, we also performed standard assays of restriction fragment length polymorphisms (RFLPs). Evaluation of 13 selected cases indicated mixed chimerism (4), complete chimerism (5), recurrence of leukemia (2), and endogenous repopulation of hematopoiesis (2) after marrow transplantation. Sensitivity of the method was determined by mixing various proportions of recipient and donor DNA; the limit of detection of the minor component in a mixture was 0.1%. PCR data correlated with RFLP data in all cases except two in which PCR proved more sensitive than RFLP. PCR amplification of VNTRs combined with oligonucleotide hybridization is a novel technique for documenting posttransplant chimerism and has advantages over RFLP analysis: high sensitivity, use of small amounts of DNA (250 ng), ease of preparation of DNA, elimination of need for restriction enzymes, and the ability to complete studies in 2 days.

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Application of long-distance polymerase chain reaction to detection of junctional sequences created by chromosomal translocation in mature B- cell neoplasms

Blood ◽

10.1182/blood.v88.3.985.985 ◽

1996 ◽

Vol 88 (3) ◽

pp. 985-994 ◽

Cited By ~ 7

Author(s):

T Akasaka ◽

M Muramatsu ◽

H Ohno ◽

I Miura ◽

E Tatsumi ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

B Cell ◽

Tumor Cells ◽

Chromosomal Translocation ◽

Pcr Amplification ◽

Chromosomal Translocations ◽

Chain Reaction ◽

Long Distance ◽

Neoplastic Cells ◽

Polymerase Chain

Abstract Junctional sequences created by chromosomal translocations in mature B- cell neoplasms, which involve immunoglobulin gene loci (IG) and putative proto-oncogenes on reciprocal partner chromosomes, are unique to neoplastic cells characterized by particular histological and immunological phenotypes. To establish a rapid and sensitive method to detect neoplastic cells carrying a specific chromosomal translocation, we have developed a novel strategy based on long-distance polymerase chain reaction (LD-PCR) amplification. Genomic DNA was extracted from tumor cells carrying t(14;19)(q32;q13), t(8;14)(q24;q32), t(3;22)(q27;q11), t(2;3)(p12;q27), or t(3;14)(q27;q32). Thirty-two to 35-mer oligonucleotide primer pairs were designed to be complementary to exons or flanking sequences of the BCL3, c-MYC and BCL6 oncogenes, and to IG constant region genes. LD-PCR with a newly available Taq polymerase for longer product synthesis successfully amplified fragments representing BCL3/C alpha junctional sequences for t(14;19); c-MYC/C mu, c-MYC/C gamma, and c-MYC/C alpha for t(8;14); BCL6/C lambda for t(3;22); BCL6/C kappa for t(2;3); 5′-BCL6/C mu, and 5′-BCL6/C gamma for t(3;14). In Burkitt's lymphoma/leukemia, all materials in which c- MYC rearrangements were detectable by conventional Southern blot hybridization showed positive LD-PCR amplification. The sizes of the amplified fragments varied from 1.8 kb to 12 kb, and these were specific to each material. Serial dilution of tumor cells or DNA in negative materials demonstrated a single band on agarose gel electrophoresis stained with ethidium bromide at a level of sensitivity of 10(-3), and hybridization with radioactive probe improved the level by one order of magnitude (1 cell in 10(4)), indicating that this LD- PCR approach is a sensitive technique capable of detecting minimal residual disease. Thus, the present study provided a useful tool for diagnosis and subsequent management of B-cell neoplasms characterized by specific chromosomal translocations.

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