DNA REFLECTION.

SUMMARY:

DNA is composed of
nucleotides bonded to a
sugar-phosphate backbone.
Double stranded DNA forms
a double helix structure.
The DNA double helix coils
up into compact structures
called chromosomes. Small
segments of the chromosome
that encode a single protein
are called genes.

PROJECT
IN
SCIENCE

LADY JULY DELA

CRUZ
TVL1-COOKERY

Development of a Reference Material of a Single

DNA Molecule for the Quality Control of PCR Testing
Junichi Mano, Shuko Hatano, Satoshi Futo, Junji Yoshii, Hiroki
Nakae, Shigehiro Naito, Reona Takabatake, and Kazumi Kitta*

National Food Research Institute, National Agriculture and Food Research

Organization, 2-1-12, Kannondai, Tsukuba, Ibaraki 305-8642, Japan

Fasmac Co., Ltd., 5-1-3, Midorigaoka, Atsugi, Kanagawa 243-0041, Japan

Japan Micro Array Consortium, 2-4-10, Kojimachi, Chiyoda-ku, Tokyo 1020083, Japan
Anal. Chem., 2014, 86 (17), pp 86218627
DOI: 10.1021/ac501314s
Publication Date (Web): July 25, 2014
Copyright 2014 American Chemical Society
*Tel: +81-29-838-7369; fax: +81-29-838-7369; e-mail: kaz@affrc.go.jp.
ABSTRACT:

We developed a reference material of a single DNA

molecule with a specific nucleotide sequence. The
double-strand linear DNA which has PCR target
sequences at the both ends was prepared as a
reference DNA molecule, and we named the PCR
targets on each side as confirmation sequence and
standard sequence. The highly diluted solution of
the reference molecule was dispensed into 96 wells
of a plastic PCR plate to make the average number
of molecules in a well below one. Subsequently,
the presence or absence of the reference molecule
in each well was checked by real-time PCR
targeting for the confirmation sequence. After an
enzymatic treatment of the reaction mixture in the
positive wells for the digestion of PCR products, the
resultant solution was used as the reference
material of a single DNA molecule with the

standard sequence. PCR analyses revealed that the

prepared samples included only one reference
molecule with high probability. The single-molecule
reference material developed in this study will be
useful for the absolute evaluation of a detection
limit of PCR-based testing methods, the quality
control of PCR analyses, performance evaluations
of PCR reagents and instruments, and the
preparation of an accurate calibration curve for
real-time PCR quantitation.

Introduction:

WHAT IS DNA?
Deoxyribonucleic acid, more commonly known
as DNA, is a complex molecule that contains all of the
information necessary to build and maintain an organism.
All living things have DNA within their cells. In fact, nearly
every cell in a multicellular organism possesses the full
set of DNA required for that organism.
However, DNA does more than specify the structure and
function of living things it also serves as the primary
unit of heredity in organisms of all types. In other words,
whenever organisms reproduce, a portion of their DNA is
passed along to their offspring. This transmission of all or
part of an organism's DNA helps ensure a certain level of
continuity from one generation to the next, while still
allowing for slight changes that contribute to the diversity
of life.

But what, exactly, is DNA? What smaller elements make

up this complex molecule, how are these elements
arranged, and how is information extracted from them?
This unit answers each of these questions, and it also
provides a basic overview of the process of DNA
discovery.