scholarly journals A Pipeline with Multiplex Reverse Transcription Polymerase Chain Reaction and Microarray for Screening of Chromosomal Translocations in Leukemia

2013 ◽  
Vol 2013 ◽  
pp. 1-13 ◽  
Author(s):  
Fei-Fei Xiong ◽  
Ben-Shang Li ◽  
Chun-Xiu Zhang ◽  
Hui Xiong ◽  
Shu-Hong Shen ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Fusion Gene ◽  
Chromosomal Rearrangements ◽  
Pcr Primers ◽  
Fusion Genes ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Set Up

Chromosome rearrangements and fusion genes present major portion of leukemogenesis and contribute to leukemic subtypes. It is practical and helpful to detect the fusion genes in clinic diagnosis of leukemia. Present application of reverse transcription polymerase chain reaction (RT-PCR) method to detect the fusion gene transcripts is effective, but time- and labor-consuming. To set up a simple and rapid system, we established a method that combined multiplex RT-PCR and microarray. We selected 15 clinically most frequently observed chromosomal rearrangements generating more than 50 fusion gene variants. Chimeric reverse primers and chimeric PCR primers containing both gene-specific and universal sequences were applied in the procedure of multiplex RT-PCR, and then the PCR products hybridized with a designed microarray. With this approach, among 200 clinic samples, 63 samples were detected to have gene rearrangements. All the detected fusion genes positive and negative were validated with RT-PCR and Sanger sequencing. Our data suggested that the RT-PCR-microarray pipeline could screen 15 partner gene pairs simultaneously at the same accuracy of the fusion gene detection with regular RT-PCR. The pipeline showed effectiveness in multiple fusion genes screening in clinic samples.

Multiplex Reverse Transcription-Polymerase Chain Reaction for Simultaneous Screening of 29 Translocations and Chromosomal Aberrations in Acute Leukemia

Blood ◽  
1998 ◽  
Vol 92 (2) ◽  
pp. 574-588 ◽  
Author(s):  
Niels Pallisgaard ◽  
Peter Hokland ◽  
Dorthe C. Riishøj ◽  
Bent Pedersen ◽  
Poul Jørgensen
Keyword(s):  
Polymerase Chain Reaction ◽  
Acute Leukemia ◽  
Chromosomal Aberrations ◽  
Reverse Transcription ◽  
Cytogenetic Analysis ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Lymphoid Leukemia ◽  
Polymerase Chain ◽  
Set Up

Abstract We have developed a multiplex reverse transcription-polymerase chain reaction (RT-PCR) reaction, which enables us to detect 29 translocations/chromosomal aberrations in patients with acute lymphoid leukemia (ALL) and acute myeloid leukemia (AML). Through the construction and optimization of specific primers for each translocation, we have been able to reduce the set-up to 8 parallel multiplex PCR reactions, thus greatly decreasing the amount of work and reagents. We show the value of our set-up in a retrospective analysis on cryopreserved material from 102 AML and 62 ALL patients. The multiplex RT-PCR detected a hybrid mRNA resulting from a structural chromosomal aberration in 45 of 102 (44%) of the AML and in 28 of 62 (45%) of the pediatric ALL cases. Importantly, in 33% of AML and in 47% of the ALL cases with cytogenetic data, submicroscopic chromosomal aberrations or masked translocations were shown that were not detected in the cytogenetic analysis either for structural reasons or because of an insufficient number of metaphases obtained. This multiplex RT-PCR system, which can handle up to 10 patients with a response time of 2 working days, is thus an important tool that complements cytogenetic analysis in the up-front screening of acute leukemia patients and should provide a rapid and efficient characterization of leukemia cells, even in situations with sparse patient material.

scholarly journals Analysis and Forecasting of Global RT-PCR Primers for SARS-CoV-2

2020 ◽  
Author(s):  
Gowri Nayar ◽  
Ed Seabolt ◽  
Mark Kunitomi ◽  
Akshay Agarwal ◽  
Kristen L. Beck ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Animal Cell ◽  
Geographic Location ◽  
High Specificity ◽  
Pcr Primers ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Over Time

Rapid tests for active SARS-CoV-2 infections rely on reverse transcription polymerase chain reaction (RT-PCR). RT-PCR uses reverse transcription of RNA into complementary DNA (cDNA) and amplification of specific DNA (primer and probe) targets using polymerase chain reaction (PCR). The technology makes rapid and specific identification of the virus possible based on sequence homology of nucleic acid sequence and is much faster than tissue culture or animal cell models. However, the technique can lose sensitivity over time as the virus evolves and the target sequences diverge from the selective primer sequences. Different primer sequences have been adopted in different geographic regions. As we rely on these existing RT-PCR primers to track and manage the spread of the Coronavirus, it is imperative to understand how SARS-CoV-2 mutations, over time and geographically, diverge from existing primers used today. In this study, we analyze the performance of the SARS-CoV-2 primers in use today by measuring the number of mismatches between primer sequence and genome targets over time and spatially. We find that there is a growing number of mismatches, an increase by 2% per month, as well as a high specificity of virus based on geographic location.

Multiplex Reverse Transcription-Polymerase Chain Reaction for Simultaneous Screening of 29 Translocations and Chromosomal Aberrations in Acute Leukemia

Blood ◽  
1998 ◽  
Vol 92 (2) ◽  
pp. 574-588 ◽  
Author(s):  
Niels Pallisgaard ◽  
Peter Hokland ◽  
Dorthe C. Riishøj ◽  
Bent Pedersen ◽  
Poul Jørgensen
Keyword(s):  
Polymerase Chain Reaction ◽  
Acute Leukemia ◽  
Chromosomal Aberrations ◽  
Reverse Transcription ◽  
Cytogenetic Analysis ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Lymphoid Leukemia ◽  
Polymerase Chain ◽  
Set Up

We have developed a multiplex reverse transcription-polymerase chain reaction (RT-PCR) reaction, which enables us to detect 29 translocations/chromosomal aberrations in patients with acute lymphoid leukemia (ALL) and acute myeloid leukemia (AML). Through the construction and optimization of specific primers for each translocation, we have been able to reduce the set-up to 8 parallel multiplex PCR reactions, thus greatly decreasing the amount of work and reagents. We show the value of our set-up in a retrospective analysis on cryopreserved material from 102 AML and 62 ALL patients. The multiplex RT-PCR detected a hybrid mRNA resulting from a structural chromosomal aberration in 45 of 102 (44%) of the AML and in 28 of 62 (45%) of the pediatric ALL cases. Importantly, in 33% of AML and in 47% of the ALL cases with cytogenetic data, submicroscopic chromosomal aberrations or masked translocations were shown that were not detected in the cytogenetic analysis either for structural reasons or because of an insufficient number of metaphases obtained. This multiplex RT-PCR system, which can handle up to 10 patients with a response time of 2 working days, is thus an important tool that complements cytogenetic analysis in the up-front screening of acute leukemia patients and should provide a rapid and efficient characterization of leukemia cells, even in situations with sparse patient material.

scholarly journals Analysis and forecasting of global real time RT-PCR primers and probes for SARS-CoV-2

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Gowri Nayar ◽  
Edward E. Seabolt ◽  
Mark Kunitomi ◽  
Akshay Agarwal ◽  
Kristen L. Beck ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Animal Cell ◽  
Geographic Location ◽  
High Specificity ◽  
Pcr Primers ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Over Time

AbstractRapid tests for active SARS-CoV-2 infections rely on reverse transcription polymerase chain reaction (RT-PCR). RT-PCR uses reverse transcription of RNA into complementary DNA (cDNA) and amplification of specific DNA (primer and probe) targets using polymerase chain reaction (PCR). The technology makes rapid and specific identification of the virus possible based on sequence homology of nucleic acid sequence and is much faster than tissue culture or animal cell models. However the technique can lose sensitivity over time as the virus evolves and the target sequences diverge from the selective primer sequences. Different primer sequences have been adopted in different geographic regions. As we rely on these existing RT-PCR primers to track and manage the spread of the Coronavirus, it is imperative to understand how SARS-CoV-2 mutations, over time and geographically, diverge from existing primers used today. In this study, we analyze the performance of the SARS-CoV-2 primers in use today by measuring the number of mismatches between primer sequence and genome targets over time and spatially. We find that there is a growing number of mismatches, an increase by 2% per month, as well as a high specificity of virus based on geographic location.

scholarly journals Analysis and Forecasting of Global RT-PCR Primers for SARS-CoV-2

2021 ◽  
Author(s):  
Gowri Nayar ◽  
Edward Seabolt ◽  
Mark Kunitomi ◽  
Akshay Agarwal ◽  
Kristen Beck ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Animal Cell ◽  
Geographic Location ◽  
High Specificity ◽  
Pcr Primers ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Over Time

Abstract Rapid tests for active SARS-CoV-2 infections rely on reverse transcription polymerase chain reaction (RT-PCR). RT-PCR uses reverse transcription of RNA into complementary DNA (cDNA) and amplification of specific DNA (primer and probe) targets using polymerase chain reaction (PCR). The technology makes rapid and specific identification of the virus possible based on sequence homology of nucleic acid sequence and is much faster than tissue culture or animal cell models. However the technique can lose sensitivity over time as the virus evolves and the target sequences diverge from the selective primer sequences. Different primer sequences have been adopted in different geographic regions. As we rely on these existing RT-PCR primers to track and manage the spread of the Coronavirus, it is imperative to understand how SARS-CoV-2 mutations, over time and geographically, diverge from existing primers used today. In this study, we analyze the performance of the SARS-CoV-2 primers in use today by measuring the number of mismatches between primer sequence and genome targets over time and spatially. We find that there is a growing number of mismatches, an increase by 2% per month, as well as a high specificity of virus based on geographic location.

1504: Molecular Diagnosis and Staging of Prostate Cancer Using Clusterin in Real Time Reverse Transcription Polymerase Chain Reaction (RT-PCR)

2006 ◽  
Vol 175 (4S) ◽  
pp. 485-486
Author(s):  
Sabarinath B. Nair ◽  
Christodoulos Pipinikas ◽  
Roger Kirby ◽  
Nick Carter ◽  
Christiane Fenske
Keyword(s):  
Prostate Cancer ◽  
Polymerase Chain Reaction ◽  
Real Time ◽  
Molecular Diagnosis ◽  
Reverse Transcription ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

Expression of hyaluronan synthase-3 in porcine oviducal epithelium during oestrus

2003 ◽  
Vol 15 (2) ◽  
pp. 99 ◽  
Author(s):  
Paisan Tienthai ◽  
Naoko Kimura ◽  
Paraskevi Heldin ◽  
Eimei Sato ◽  
Heriberto Rodriguez-Martinez
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Hyaluronan Synthase ◽  
Critical Periods ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Mean Values ◽  
Polymerase Chain ◽  
Sperm Reservoir ◽  
Oviduct Fluid

Hyaluronan (HA) has been related to fertilization and embryo development in the pig. Furthermore, HA is present in pig oviduct fluid and the lining epithelium, particularly of the pre-ovulatory sperm reservoir. Because the mechanisms that regulate HA synthesis have not yet been clarified, semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) was conducted to assess the expression of mRNAs of two HA-synthesizing enzymes (has2 and has3) in the oviduct epithelium (uterotubal junction, isthmus, ampullary–isthmic junction and ampulla segments) of non-inseminated (control) and inseminated (treatment) sows at pre-, peri- and post-ovulatory oestrus. Only has3 mRNA was detected; it was present in all tubal segments of both control and treatment samples. The level of has3 expression did not vary significantly between non-inseminated and inseminated specimens, but there was a tendency (NS) for increased mean values during the peri- and post-ovulatory stages compared with pre-ovulation. It is concluded that has3 is expressed by the porcine endosalpinx epithelium and the levels of expression do not vary during the critical periods of sperm transport and fertilization, despite fluctuating levels of HA in the tubal fluid at corresponding periods.

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