TY - JOUR
T1 - Simultaneously extracting DNA, RNA, and protein using kits
T2 - Is sample quantity or quality prejudiced?
AU - Mathieson, William
AU - Thomas, Gerry A.
PY - 2013/2/1
Y1 - 2013/2/1
N2 - Interdisciplinary "omics" research and the stringent quality requirements of array-based technologies require the simultaneous yet efficient extraction of DNA, RNA, and protein from the same tissue block. However, the few commercially available simultaneous extraction kits have not been evaluated. We compare the TriplePrep (GE Healthcare) and AllPrep (Qiagen) kits using good, intermediate, and poor quality tissue with specialist single-extract methods: Puregene (DNA), RNeasy (RNA), and homogenizations into buffer (protein). The following parameters were evaluated: DNA - yield (total DNA and double-stranded), purity (260:280 and 260:230), and integrity (gel electrophoresis); RNA - yield, purity, and integrity (RNA integrity numbers [RINs] and quantitative reverse transcription polymerase chain reaction [Q-RT-PCR]); protein - yield and quality (two-dimensional difference gel electrophoresis [2D-DIGE]). Puregene DNA yields were 183% and 506% those of TriplePrep and AllPrep, respectively. For RNA, AllPrep and RNeasy were indistinguishable, but their yields were 412% to 588% those of TriplePrep (depending on block condition) and their between-sample variability was better. TriplePrep protein yields were 57% those of the control, and 6.9% of the gel spots were more than 2-fold altered. However, AllPrep yields were 20% of the control, with 11% of the gel spots being more than 2-fold altered. Therefore, TriplePrep outperformed AllPrep in DNA and protein extractions, the reverse was true for RNA, but neither kit achieved optimal efficiency because both yield and quality were compromised.
AB - Interdisciplinary "omics" research and the stringent quality requirements of array-based technologies require the simultaneous yet efficient extraction of DNA, RNA, and protein from the same tissue block. However, the few commercially available simultaneous extraction kits have not been evaluated. We compare the TriplePrep (GE Healthcare) and AllPrep (Qiagen) kits using good, intermediate, and poor quality tissue with specialist single-extract methods: Puregene (DNA), RNeasy (RNA), and homogenizations into buffer (protein). The following parameters were evaluated: DNA - yield (total DNA and double-stranded), purity (260:280 and 260:230), and integrity (gel electrophoresis); RNA - yield, purity, and integrity (RNA integrity numbers [RINs] and quantitative reverse transcription polymerase chain reaction [Q-RT-PCR]); protein - yield and quality (two-dimensional difference gel electrophoresis [2D-DIGE]). Puregene DNA yields were 183% and 506% those of TriplePrep and AllPrep, respectively. For RNA, AllPrep and RNeasy were indistinguishable, but their yields were 412% to 588% those of TriplePrep (depending on block condition) and their between-sample variability was better. TriplePrep protein yields were 57% those of the control, and 6.9% of the gel spots were more than 2-fold altered. However, AllPrep yields were 20% of the control, with 11% of the gel spots being more than 2-fold altered. Therefore, TriplePrep outperformed AllPrep in DNA and protein extractions, the reverse was true for RNA, but neither kit achieved optimal efficiency because both yield and quality were compromised.
KW - AllPrep
KW - DNA
KW - Extraction
KW - Protein
KW - Puregene
KW - RNA
KW - RNeasy
KW - Simultaneous
KW - TriplePrep
UR - http://www.scopus.com/inward/record.url?scp=84869124886&partnerID=8YFLogxK
U2 - 10.1016/j.ab.2012.10.006
DO - 10.1016/j.ab.2012.10.006
M3 - Article
C2 - 23068038
AN - SCOPUS:84869124886
SN - 0003-2697
VL - 433
SP - 10
EP - 18
JO - Analytical Biochemistry
JF - Analytical Biochemistry
IS - 1
ER -