[Research frontier] CRISPR Detection Trends
The
nucleic acid
testing for quick
specific
ity
detection of infectious
pathogens is
widely
recognized, such as
P
olymerase
C
hain
R
eaction (PCR)
. However
it is expensive
due to the
expensive equipment
and
the
highly trained Researchers
.In this way,
CRISPR-Cas
system have recently emerged as an alternative to PCR-based diagnosis. CRISPR diagnosis is also known as next-generation molecular diagnostic technology, which mainly utilizes RNA to detect biomarkers in biological fluids
(
such as blood or urine
). By processing the
sample
preparation
(including pre-amplification), target recognition, signal release
and
testing, t
he CRISPR/Cas system
become the
potential diagnostic tool for disease and pathogen detection
. It is
reported
that
CRISPR/Cas biosensing systems have the advantages of high resolution, high sensitivity, low cost,
simple
operation
and
storage.
In the following,
we
select and share the latest cases of CRISPR detection applications in humans, animals, and plants to provide some ideas and help for CRISPR detection research.
Colorimetric Detection of Pseudomonas aeruginosa
Pseudomonas aeruginosa(
P. aeruginosa), a Gram-negative pathogen, has attracted increasing attention in post-operative infections and poses a new threat to human health.
The
P. aeruginosa
is
difficult to
be
control
led becasue of the
wide range of habitats
(
such as water, air, animals, and humans
). Developing a
reliable
detecion methods for
P. aeruginosa
is
particularly
important but still a huge challenge.
P. aeruginosa causes long-term chronic diseases by causing bone and joint infections in patients with compromised immune function. In this study, assist
ed
by
target
identification, the
self-primers initiate
d
the production of single-stranded DNA, which
can be
recognized by CRISPR-Cas12a. Therefore, the trans-cleavage activity of Cas12a enzyme
wa
s activated, digesting the silver ion-chelating aptamer sequence. Due to the strong trans-cleavage activity of Cas12a enzyme,
the
DNA aptamer-based silver ions detection
exhibits higher sensitivity.
Besides, the
limit of detection
(LOD)
in this
method is 21 cfu/mL, which
enables a
broad application in early diagnosis of infection.
Schematic diagram of colorimetric method based on CRISPR-Cas12a
Reference: Hu, Jiangchun & Liang, Ling & He, Mingfang & Lu, Yongping. (2023). Sensitive and Direct Analysis of Pseudomonas aeruginosa through Self-Primer-Assisted Chain Extension and CRISPR-Cas12a-Based Color Reaction. ACS Omega. 8. 10.1021/acsomega.3c04180.
- E DITGENE provides all the CRISPR raw materials used in the research , such as Cas12a, Cas12b, Cas13a, RPA, reporter . Click to view the list of CRISPR detection raw materials
Developing a Rapid Detection Method for Plant Pathogens
Fusarium head blight (FHB) is a global cereal disease caused by species of Fusarium. Both Fusarium graminearum and Fusarium asiaticum are the causal agents of FHB in China. Asian grass is a dominant species in the middle and lower reaches of the Yangtze River (MLRYR) and southwest China . Therefore, timely detection of Fusarium asiaticum is crucial to controlling the disease and preventing mycotoxins from entering the food chain. T his study optimized reaction conditions based on CRISPR Cas12a technology and developed a rapid, sensitive, and cost-effective detection method for Fusarium asiaticum. The optimized method showed specificity in detecting Fusarium asiaticum without detecting any other 14 strains of Fusarium and 3 non-Fusarium species. The limit of detection of this method is 20 ag/µL, and even Fusarium asiaticum in corn and wheat grains can be diagnose d after 3 - day inoculation. This method truly provide an efficient and robust detection platform for accelerating the detection .
The detection process of the developed RPA-Cas12a-LFD detection system
Reference: Zhang, J.; Liang, X.; Zhang, H.; Ishfaq, S.; Xi, K.; Zhou, X.; Yang, X.; Guo, W. Rapid and Sensitive Detection of Toxigenic Fusarium asiaticum Integrating Recombinase Polymerase Amplification, CRISPR/Cas12a, and Lateral Flow Techniques. Int. J. Mol. Sci. 2023 , 24 , 14134. https://doi.org/10.3390/ijms241814134
- If you also want to utilize CRISPR detection to diagnosis pathogens , E DITGENE offers a complete solution,which is with high sensitiv ity , quick detect ion and strong specific ity. For more information, please visit us .
Detection and Serotyping Identification of Streptococcus suis Serotype 2
Streptococcus suis Serotype 2
is an e
conomically important
zoonotic pathogen that can cause sepsis, arthritis, and meningitis in pigs and humans.
Streptococcus suis Serotype 2
may
cause significant economic losses to the pig industry
,
threat
ening
t
he
public health. Therefore, developing an accurate and rapid detection method is significan
t
for epidemic prevention and control.
This study developed a detection and serotyping platform
named
Cards-SSJ/K
for serotype 2 of swine streptococcus based on recombinant polymerase amplification technology (RPA) and CRISPR-Cas12a system. The detection method has a detection limit of 10 CFU and a detection time of less than 60 minutes. It has no cross-reactivit
ion
with other
S
erotypes of
S
wine
S
treptococcus, closely related
S
treptococcus or
other
common pig pathogens
.
and Cards-SSK can distinguish between serotype 2 and serotype 1/2. The results of
Cards-SSJ
and qPCR detection of
S
wine
S
treptococcus serotype 2 are the same.
The results
showed that although the reagent cost of Cards-SSJ is relatively high compared to PCR and qPCR, it takes less time and has lower requirements for equipment and personnel. Therefore, it is an excellent method for POCT detection of
S
wine
S
treptococcus serotype 2.
The process of detecting serotype 2 of S wine S treptococcus based on CRISPR Cas12a technology
Refernce: Wang, Lu & Sun, Jing & Zhao, Jiyu & Bai, JieYu & Zhang, Yueling & zhu, Yao & Zhang, Wanjiang & Wang, Chunlai & Langford, Paul & Liu, Siguo & li, Gang. (2023). A CRISPR-Cas12a-based platform facilitates the detection and serotyping of Streptococcus suis serotype 2. Talanta. 267. 125202. 10.1016/j.talanta.2023.125202.
- E DITGENE is specialized on developing and optimizing CRISPR/Cas-based genome editing technologies. The technical team has obtained Cas12 and Cas13 gen ome editing protein with better-perform ance through the self-developed Fish-bait purification process , and developed a unique crRNA design system . This has led to the establishment of the FASST detection technology, which greatly improves the cleavage activity of Cas enzyme and CRISPR detection sensitivity.
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Test of LwaCas13a Activity
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