CRISPR/Cas9 - regulatory issues

In the previous blog entry the basics of the CRISPR/Cas9 method has been elucidated. Now we want to talk about the future of this method. However to talk about the future of CRISPR/Cas9 we have to talk about the legal regulations concerning the application of this method. The following blog entry considers the contemporary juridical situation and will give ideas on how to deal with that issue in the near future.

To regulate the application and the admission of CRISPR/Cas9 it is of great importance to determine whether CRISPR/Cas9 treated organism are to be seen as GMOs (genetically modified organisms) or not. In Switzerland, the Federal Office for the Environment has submitted a substantial study concerning the classification of GMOs in the year 2012. However in this study called “Grundlagen für die Klärung offener Fragen bei der rechtlichen Regulierung neuer Pflanzenzuchtverfahren” (Vogel, 2012) the CRISPR/Cas9 method is not mentioned. This shows that the topic is still at issue and is debated in Switzerland as well as in whole Europe (Lucht, 2016).

In 2015, the “Swiss Expert Comitee for Biosafety SECB” proposed to not classify the products of CRISPR/Cas9 as GMOs because the genetical modifications used for the production of these organism can’t be detected anymore in the finished products (SECB, 2015).

The “answer” to this Proposal by the SECB, followed in march 2016 by the “Federal Ethics Comitee on Non-Human Biotechnology ECNH”. The Comitee concludes in the related paper, that there is not enough knowledge on this topic at this time to evaluate if the concerning organisms have to be treated as GMOs or not. This is why the ECNH postulates to act by the “precautionary principle” (ECNH, 2016).

As a last point there is to say that this debate concerning the new techniques of plantbreeding, is held almost exclusively among experts at this time. This is probably the reason why there isn’t a distinctive public opinion on this topic yet. However it is beyond discussions that the research on this issue has to continue and that the populace has to be integrated in this debate as well.

References:

Vogel B. (2012). Neue Pflanzenzuchtverfahren - Grundlagen für die Klärung offener Fragen bei der rechtlichen Regulierung neuer Pflanzenzuchtverfahren. Baudirektion des Kantons Zürich, AWEL, SBS

Lucht J. (2016). Fact Sheet – Neue gentechnische Verfahren. scienceindustries Zürich

Swiss Expert Comitee for Biosafety SECB (2015). Bericht der EFBS zu Neuen Pflanzenzuchtverfahren. SECB

Federal Ethics Comitee on Non-Human Biotechnology ECNH (2016). Neue Pflanzenzüchtungsverfahren – ethische Überlegungen. ECNH

CRISPR/Cas9

Genome engineering, in the colloquial speech often denoted as “gene manipulation” is a politically, socially and economically lively debated topic with proponents and opponents coming from different lobbies. It is undisputed that Technologies for making and manipulating DNA have enabled advances of great importance. One of these technologies, the CRISPR/Cas9 is highly topical as it has been declared by the Science-magazine as the „Breakthrough oft the Year 2015“.

In the mid 2000s, scientists discovered so called CRISPRs (clustered regularly interspaced short palindromic repeats) in the genome of Escherichia coli. Later on, CRISPRs have been detected in various species of bacteria and archae (Doudna & Charpentier, 2012). These sequences are part of the CRISPR/Cas system which acts as a protection mechanism against viral invasion. CRISPR-associated proteins (Cas) are able to bind and specific RNA sequences , the so called crRNA-repeat sequence. This sequence is followed by a crRNA-spacer sequence which is determining for its binding on the tareted DNA sequence. In turn, the enzyme endonuclease Cas9 cuts this DNA in its close environment. With this ability, the CRISPR/Cas system is able to find, cut and destroy viral DNA.

Due to the the ability of this system to edit specific sequences of the DNA with a high precision it can be used as a mechanism to edit eukaryotic DNA as well, as J. Doudna and E. Charpentier discovered in 2012. The simplicity of this system makes it highly interesting as it only requires three components (crRNA, trRNA and tha Cas9 enzyme).

Meanwhile, the CRISPR/Cas9 system has been successfully used to target important genes in a broad variety of species including plants and humans (Reis, 2014). Presumably the targeting and editing of genes in plants of agronomic interests is going to be the most important application area. For example the team around Dr. Cristobal Uauy tried to target a gene coding for a plasma membrane protein which is thought to be involved in grain dormancy.

Despite of all the advantages this type of technology definitely has, its usage is to handle with care. Concerning the near future of this technology, the focus should lay on a suggestive regulation on the application of such a biochemical “superweapon”.

References:

Doudna, J.A. & Charpentier, E. (2014). The new frontier of genome engineering with CRISPR-Cas9.

http://blogs.biomedcentral.com/on-biology/2015/11/30/using-crisprcas9-genetically-modify-crops/


https://www.neb.com/tools-and-resources/feature-articles/crispr-cas9-and-targeted-genome-editing-a-new-era-in-molecular-biology

Micropropagation of the "Shidareguwa" mulberry

With its graceful canopy and its several application possibilities in the food-, cosmetical- and pharmaceutical industry, the cultivation of the “Shidareguwa” mulberry (Morus alba var. Shidareguwa) is a matter of public and economic interest. Due to the difficult propagation of Morus alba by using conventional methods, the in vitro micropropagation of M. alba gains in importance. The following review is about the paper “Micropropagation of a difficult-to-root weeping mulberry (Morus alba var. Shidareguwa): A popular variety for ornamental purposes” (Aroonpong, Chang, 2015).

-       The study examined the application of existing in vitro propagation methods on Morus alba. With its potential economic benefits as a driving force, the development of an efficient and cost-effective propagation Method is of great value and would improve and accelerate the cultivation of Morus alba.

-       Either shoot tips or nodes of different ages (Node 1,2 & 3) have been used as explants and were cut off from a 3-year old field-grown Morus alba. These explants have been disinfected and afterwards put in a MS medium with variable content of the phytohormon BAP. For the rooting the medium has been changed to an MS medium with different concentrations of the auxine IBA. To be able to compare the rooting ability, explants of Morus alba have been cultured ex vitro as well, after being treated with IBA.

-       Survival, exo- and endophytic contaminations and browning of the explants varied depending on the used part of the plants as showed in Figure 2 in the publication (p. 322). The shoot tips showed the highest survival rate (100%) whereas the nodes showed decreasing survival rates with increasing age of the nodes. Nodes of increasing age also showed increasing endo- and exophytic contaminants probably due to the specificity of the vascular tissues which are less specificated in the shoot tips and the younger nodes.

-       The success in the acclimatisation of the in vitro culture is depending mainly on the duration of the acclimatisation process. It takes more time to acclimatise plants coming from an in vitro culture compared to plants propagated ex vitro. Also the amount of leaves and the consequential better uptake of carbon energy seems to accelerate the rooting process during the acclimatisation stage.