Design and Implementation of Polymerase Chain Reaction Device for Aptamers Selection of Tumor Cells

2020 ◽  
Vol 20 (3) ◽  
pp. 1332-1340
Author(s):  
Chao Wang ◽  
Fan Meng ◽  
Yanping Huang ◽  
Nongyue He ◽  
Zhu Chen
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Tumor Cells ◽  
Pcr Amplification ◽  
Chain Reaction ◽  
Control Module ◽  
Aptamer Selection ◽  
Nucleic Acid Aptamers ◽  
Polymerase Chain ◽  
Selection Of

Nucleic acid aptamers are a kind of one-dimensional biological nanomaterials and have found many applications. This paper designed and implemented a polymerase chain reaction (PCR) amplification device with a reaction volume of 500 μL, which can be used for the amplification of nucleic acid aptamers of tumor cells in the aptamer selection. This device mainly includes a control module, a temperature measuring module, a PCR amplification tube, a metal tank module, a liquid crystal display (LCD) and operation module and a cooling module. The new PCR amplification chamber is matched with the designed metal tank to ensure the temperature uniformity of the PCR amplification solution. The control module based on the STM32F103RCT6 manages the workflow of the entire device. The PCR amplification chamber and PT100 sensors on the metal tank formed a closed-loop feedback system, and the incremental proportional-integral-derivative (PID) algorithm was used to achieve the precise temperature control. In addition, we introduced the Smith predictive compensation algorithm to solve the temperature hysteresis problem of the PCR amplification chamber. The experimental results showed that the PCR device can meet the requirements for the nucleic acid aptamer selection of tumor cells. The device can also be used in other experiments with large-volume PCR amplification.

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

Blood ◽  
1996 ◽  
Vol 88 (3) ◽  
pp. 985-994 ◽  
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.

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

Blood ◽  
1996 ◽  
Vol 88 (3) ◽  
pp. 985-994 ◽  
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

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.

scholarly journals Easy and Efficient DNA Extraction from Woody Plants for the Detection of Phytoplasmas by Polymerase Chain Reaction

Plant Disease ◽  
1999 ◽  
Vol 83 (5) ◽  
pp. 482-485 ◽  
Author(s):  
Margaret J. Green ◽  
Dan A. Thompson ◽  
Donald J. MacKenzie
Keyword(s):  
Polymerase Chain Reaction ◽  
Plant Material ◽  
Woody Plant ◽  
Leaf Roll ◽  
Chain Reaction ◽  
Efficient Procedure ◽  
Identical Manner ◽  
Total Dna ◽  
Polymerase Chain ◽  
Selection Of

A simple and efficient procedure for the extraction of high-quality DNA from phytoplasma-infected woody and herbaceous plants for polymerase chain reaction (PCR) detection is described. This procedure does not require phenol, chloroform, or alcohol for the precipitation of nucleic acids. Herbaceous and woody plant material are extracted in an identical manner with no additional purification or enrichment steps required. The method utilizes commercially available microspin-column matrices, and the extraction of total DNA can be achieved in less than 1 h. The method has been used to successfully purify phytoplasma DNA from whole leaves, leaf petioles and midribs, roots, and dormant wood from a diverse selection of plant material. The phytoplasmas detected by PCR include pear decline, western X-disease, peach yellow leaf roll, peach rosette, apple proliferation, Australian grapevine yellows, and Vaccinium witches'-broom.

scholarly journals Amplification of human beta-actin gene by the reverse transcriptase-polymerase chain reaction: implications for assessment of RNA from formalin-fixed, paraffin-embedded material.

1996 ◽  
Vol 44 (10) ◽  
pp. 1205-1207 ◽  
Author(s):  
A Dakhama ◽  
V Macek ◽  
J C Hogg ◽  
R G Hegele
Keyword(s):  
Polymerase Chain Reaction ◽  
Pcr Amplification ◽  
Housekeeping Gene ◽  
Chain Reaction ◽  
Viral Genomes ◽  
Negative Results ◽  
Beta Actin ◽  
Target Nucleic Acid ◽  
Formalin Fixed Paraffin Embedded ◽  
Polymerase Chain

The polymerase chain reaction (PCR) is a powerful method that allows enzymatic amplification of rate target nucleic acid sequences. It has been applied to the amplification of viral genomes from paraffin-embedded pathology specimens. However, interpretation of negative results requires amplification of a housekeeping gene such as beta-actin. In the present study we used specific oligonucleotide primers previously designed to amplify both the genomic DNA and the mRNA transcript from paraffin-embedded tissue. These products have predicted sizes of 250 BP and 154 BP, respectively, but our results showed that PCR amplification only (without reverse transcription) unexpectedly generated the 154-BP product. Further investigation of the nature of this product demonstrated that it originated from the amplification of DNA, not RNA. We conclude that the 154-BP product generated by these primers cannot be exclusively considered as beta-actin RNA product and should not be used to assess successful extraction of RNA, to ascertain its integrity, or to normalize for the total amount of RNA assayed by RT-PCR from paraffin-embedded tissue.

scholarly journals A Self-Contained Portable Polymerase-Chain-Reaction System Integrated with Electromagnetic Mini-Actuators for Bi-Directional Fluid Transport

2011 ◽  
Vol 27 (3) ◽  
pp. 357-364
Author(s):  
B. T. Chia ◽  
S.-A. Yang ◽  
M.-Y. Cheng ◽  
C.-W. Lin ◽  
Y.-J. Yang
Keyword(s):  
Polymerase Chain Reaction ◽  
Fluid Transport ◽  
Point Of Care ◽  
Reaction System ◽  
Thermal Characteristics ◽  
Pcr Amplification ◽  
Chain Reaction ◽  
Rapid Temperature ◽  
Small Footprint ◽  
Polymerase Chain

ABSTRACTIn this paper, the development of a portable polymerase chain reaction (PCR) device is presented. Integrating electromagnetic mini-actuators for bi-directional fluid transport, the proposed device, whose dimension is 67mm × 66mm × 25mm, can be fully operated with a 5V DC voltage. The device consists of four major parts: A disposable channel chip in which PCR mixture is manipulated and reacted, a heater chip which generates different temperature zones for PCR reaction, a linear actuator array for pumping PCR mixture, and a circuit module for controlling and driving the system. The advantages of the device include the rapid temperature responses associated with continuous-flow-type PCR devices, as well as the programmable thermal cycling associated with chamber-type PCR devices. The thermal characteristics are measured and discussed. PCR amplification is successfully performed for the 122 bp segment of MCF-7/adr cell line. Due to its small footprint, this self-contained system potentially can be employed for point-of-care (POC) applications.

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