i used the crispr/cas9 on a non-model vertebrate animal, it quite worked. I just used BLAST to align the target sequences check if it was right in this domestic animal. after, I used the following website to design my sgrna，http://cas9.cbi.pku.edu.cn/. the founder animal was born and sequenced for the target sites, it did work.
There are lots of available Crispr/Cas9 vectors suited for different oraganisms available in the Addgene site. Here is the link http://www.addgene.org/CRISPR/ , you can order it as low as 65$ which is very cheap. They provide all the necessary protocols for designing the guided RNA, plasmid infection and target efficiency.
Please, share your opinion and/or advices here in order to improve the use CRISPR/Cas9 system for functional genomics in non-model insects (but Drosophila experiences are welcome). Thank you so much again.
Two recent contributions from silkworms:
Wei W, Xin H, Roy B, Dai J, Miao Y, Gao G. Heritable genome editing with
CRISPR/Cas9 in the silkworm, Bombyx mori. PLoS One. 2014 Jul 11;9(7):e101210.
doi: 10.1371/journal.pone.0101210. eCollection 2014. PubMed PMID: 25013902;
PubMed Central PMCID: PMC4094479.
Zhang Z, Aslam AF, Liu X, Li M, Huang Y, Tan A. Functional analysis of Bombyx
Wnt1 during embryogenesis using the CRISPR/Cas9 system. J Insect Physiol. 2015
Jun 9. pii: S0022-1910(15)00123-7. doi: 10.1016/j.jinsphys.2015.06.004. [Epub
ahead of print] PubMed PMID: 26070541.
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Reading suggestion:
Chen L, Wang G, Zhu YN, Xiang H, Wang W. Advances and perspectives in the application of CRISPR/Cas9 in insects. Dongwuxue Yanjiu. 2016 Jul 18;37(4):220-8.
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Please, let's contribute with:
https://www.researchgate.net/post/How_to_design_gRNA_for_non-model_organisms
CRISPR-Cas strategy is working for model as well as non-model organisms with more method optimizations in non-model organisms. Basic questions always remains what kind of questions someone asking far? We can target single cell manipulations as well as tissue specific manipulations at certain stages of life history.
Recently few studies came up while making a comprehension in the field where one can have an alternative ways also rather than single cell stage manipulations of gene of target using CRISPR.
Non-viral/viral based tissue specific targeting of genetic manipulations examples of CRISPR success stories using lipid nanoparticles
https://www.nature.com/articles/nbt.4005
https://www.nature.com/articles/am2017185
:: The latest advances ::
Metabolic pathway interruption: CRISPR/Cas9-mediated knockout of tryptophan 2,3-dioxygenase in Tribolium castaneum.
Adrianos S, Lorenzen M, Oppert B.
J Insect Physiol. 2018 May - Jun;107:104-109. doi: 10.1016/j.jinsphys.2018.03.004. Epub 2018 Mar 15.
PMID: 29551569
New insight into the mechanism underlying the silk gland biological process by knocking out fibroin heavy chain in the silkworm.
Cui Y, Zhu Y, Lin Y, Chen L, Feng Q, Wang W, Xiang H.
BMC Genomics. 2018 Mar 26;19(1):215. doi: 10.1186/s12864-018-4602-4.
PMID: 29580211 Free PMC Article
Drosophila multicopper oxidase 3 is a potential ferroxidase involved in iron homeostasis.
Wang X, Yin S, Yang Z, Zhou B.
Biochim Biophys Acta. 2018 Aug;1862(8):1826-1834. doi: 10.1016/j.bbagen.2018.04.017. Epub 2018 Apr 21.
PMID: 29684424
Efficacy of In Vivo Electroporation on the Delivery of Molecular Agents into Aphid (Hemiptera: Aphididae) Ovarioles.
Jamison BV, Thairu MW, Hansen AK.
J Insect Sci. 2018 Mar 1;18(2). doi: 10.1093/jisesa/iey041.
PMID: 29718443 Free PMC Article
CRISPR/Cas9 mediated high efficiency knockout of the eye color gene Vermillion in Helicoverpa zea (Boddie).
Perera OP, Little NS, Pierce CA 3rd.
PLoS One. 2018 May 17;13(5):e0197567. doi: 10.1371/journal.pone.0197567. eCollection 2018.
PMID: 29771955 Free PMC Article
Genome editing reveals the function of Yorkie during the embryonic and early larval development in silkworm, Bombyx mori.
Xu X, Zhang Z, Yang Y, Huang S, Li K, He L, Zhou X.
Insect Mol Biol. 2018 May 24. doi: 10.1111/imb.12502. [Epub ahead of print]
PMID: 29797485
Translucent larval integument and flaccid paralysis caused by genome editing in a gene governing molybdenum cofactor biosynthesis in Bombyx mori.
Fujii T, Yamamoto K, Banno Y.
Insect Biochem Mol Biol. 2018 Aug;99:11-16. doi: 10.1016/j.ibmb.2018.04.008. Epub 2018 May 25.
PMID: 29803701
Identification and functional characterization of doublesex gene in the testis of Spodoptera litura.
Du Q, Wen L, Zheng SC, Bi HL, Huang YP, Feng QL, Liu L.
Insect Sci. 2018 May 29. doi: 10.1111/1744-7917.12608. [Epub ahead of print]
PMID: 29808584
BmHpo mutation induces smaller body size and late stage larval lethality in the silkworm, Bombyx mori.
Xu X, Bi HL, Zhang ZJ, Yang Y, Li K, Huang YP, Zhang Y, He L.
Insect Sci. 2018 May 29. doi: 10.1111/1744-7917.12607. [Epub ahead of print]
PMID: 29808962
Consequences of resistance evolution in a Cas9-based sex conversion-suppression gene drive for insect pest management.
KaramiNejadRanjbar M, Eckermann KN, Ahmed HMM, Sánchez C HM, Dippel S, Marshall JM, Wimmer EA.
Proc Natl Acad Sci U S A. 2018 Jun 12;115(24):6189-6194. doi: 10.1073/pnas.1713825115. Epub 2018 May 29.
PMID: 29844184
Genome editing in Bombyx mori: New opportunities for silkworm functional genomics and sericulture industry.
Ma SY, Smagghe G, Xia QY.
Insect Sci. 2018 May 29. doi: 10.1111/1744-7917.12609. [Epub ahead of print] Review.
PMID: 29845729
CRISPR/Cas9-based heritable targeted mutagenesis in Thermobia domestica: A genetic tool in an apterygote development model of wing evolution.
Ohde T, Takehana Y, Shiotsuki T, Niimi T.
Arthropod Struct Dev. 2018 Jun 13. pii: S1467-8039(18)30073-2. doi: 10.1016/j.asd.2018.06.003. [Epub ahead of print]
PMID: 29908341
INTERESTING:
Gene editing technologies and applications for insects
https://www.ncbi.nlm.nih.gov/pubmed/30551769
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