Sunday, 15 April 2018

Synthetic Biology-What can we expect...

                        Synthetic Biology-What can we expect:

Synthetic Biology deals with the study of existing biological systems and structures, redesigning them and formation of new biological systems. This field combines knowledge and apply it for the betterment of mankind from a number of fields including; genetic engineering, molecular biology, biotechnology, biophysics, molecular engineering, computer and electrical engineering.
The field of synthetic biology is continuously revolutionizing the world as it provides human with all the necessary knowledge needed for dealing with various problems, diseases and helping mankind to think beyond boundaries and enabling them to devise such techniques and technologies which were considered impossible before.

CONTRIBUTIONS:
Following are some recent contributions by the field of synthetic biology.

1.Chimeric Organisms:
                                        The word chimera is driven from Greek mythology which means a monster having head of a lion, body of a goat, and tail of a snake, ultimately a combination of various living beings. In biology such thing is termed as chimera which contains materials from multiple living sources, it could be a chimeric DNA or a chimeric organism. Now a day scientific community across the globe is working on developing chimeric organisms having human genetic material and capable of producing various human organs which could ultimately be used for clinical trials, testing of various new medicines, and for transplant purposes. At the moment the demand for organ transplant is 4 times the supply. So, successful formation of chimeric organisms having human cells and organs will be helpful to meet this requirement. Multiple experiments were carried before human trials:

   · Inserting rat stem cells in to mice forming rat-mice hybrid.
   ·  Inserting rat stem cells in to pig no positive result.
   ·  Inserting human stem cells in to pig embryo. Positive result and formation of human –pig hybrid.
Procedure for the formation of Human-Pig hybrid

2.Bio-fuels:
                  Petro-chemicals are now becoming an out dated source to power our vehicles.  Continuous CO2 emissions and global warming issues are forcing man kind to shift towards a viable source. Bio-fuels are the best possible alternative till now to power our vehicles. The formation of bio-fuels is a complex process and greatly dependent on synthetic biology. Formation of such bio-catalysts which can effectively and efficiently convert sugars in to desired products is because of molecular engineering and synthetic biology. Researchers are now struggling to develop more efficient bio-catalysts which can produce high quantity of bio-fuels out of sugars and also do it more cheaply in order to bring the cost of bio-fuels way lower than the traditional petrochemicals. Many developed and highly developing nations are already producing bio-fuels in large quantities and are using it. USA, Brazil and Germany are the top 3 producers followed by China, Argentina, France, Indonesia, Canada, Thailand, and Colombia.
Top 10 Bio-fuel Producers (Courtesy World Economic Forum)

3.Bio-sensors:
                          Bio-sensors are such organisms which are modified to give out certain signals upon completion of some physiological and biological process. These kinds of sensors can be used to detect and quantify certain reactions and biological process. These bio-sensors provide an effective way to monitor cells under in-vivo conditions helping scientists to better understand the survival, migration, and possible tumourigenicity in animal cells.

4.Data Storage:
                           Storing data is a key part of learning and evolving as it enables us to look back in our past and see where we did right or wrong and what is necessary to be done to deal with a specific problem. But to store data for a long time is a challenge. Normal hard drives have a life span of 5-10 years and are unable to store data for centuries, which forces us to make its copies regularly and if one doesn’t do it he’ll lose the data forever. Scientist are working to deal with the problem by creating such thing which can hold data for centuries without any issue. Science is now looking at nature for a solution. Researchers at The ETH Zurich, Switzerland believe that answer lies in natural data storage system of living cells: THE DNA, So compact and complex that a 1gm can theoretically store all data of TECH Giants like Google, Amazon, Facebook. In storage terms 1 gm DNA can store 455 exabytes of information and 1 exabyte is equal to a billion gigabytes.
DNA is a unique way of nature to store data for centuries in form of fossils. Scientist has succeeded in extracting genome of 110,000-year-polar bear and recently a 700,000-year old horse. Robert Grass of Dept. of Chemistry and applied Bio-science said:
       "We have found ways of making DNA very stable, So we want to combine high storage density of DNA with stability of DNA found in fossils"
Future USB's might be made up of DNA

5.Nano-particles:
                          Use of nanotech in various fields of science including biology is increasing day by day. Nanotech in biology has shown some promising results so far indicating the impact it could make on our lives. Many kinds of nano-robots are used as drug delivery systems which deliver a certain medicine directly in to the infected cells preventing the uptake of the drug by healthy cells and avoiding some malfunctioning and side effects of the drug. These nano-robots are enabling researchers to deliver the drugs in a very controlled manner and dosage. Various nano-particles are also being used to reduce anti-biotic resistance. Various nano-particles are under clinical trial for treatment of various diseases. E.g.:
   · Abraxane are the nanoparticle albumin bound paclitaxel used for the treatment of various type of cancers including breast cancer and pancreatic cancer.
   · C-dots are small silica based nano particles which give fluorescence with an organic  dye and is being used for easy detection of tumour cells by surgeons.
   · Nano particles are also used for treatment of eye diseases including cataract and dry eye disease.
Nano-robots directly delivering medicine to individual cells

6.Genome editing:
                               Every single development inside a living organism is managed by its genome all the necessary data is encoded inside the genome. A minor error in this genome can result in to severe and incurable diseases but thanks to genome editing these issues are now less complicated. Genome editing is the thing these days. Everyone across the world is talking about it and the impact it can cause on the human life. Imagine of an incurable disease which can be treated with replacing the error gene with healthy one simply by cutting the first are replacing the latter with it. According to an estimate almost 85-90% of genetic disorders are incurable but with this gene editing we can deal with those issues. Along with treatment of diseases genome editing can also be used for solving many other problems. It can be used in production of more advanced and efficient plants, animals and even humans. It can be used in plants to make them resistive against various diseases without any usage of foreign DNA, improving yield outputs of the plant, making them more adoptive to harsh environmental conditions, etc. Genome editing is an efficient way to manipulate a living being according to our desire and can prove extremely beneficial in near future. Currently the most effevtive genome editing tool is the CRISPR/CAS-9 , others include CRISPR/CPF-1, Zinc finger nuclease, and TALENS.

Sunday, 1 April 2018

Classification of CRISPR-CAS System

Classification of CRISPR-CAS System

Ever since its discovery the CRISPR CAS System has become a vital tool in the field of life sciences. What was originally discovered as bacterial immunity system in the bacteria is now being used in the field of life sciences as a broad range genome editing tool. After scientist successfully hijacked this immunity system and turned it in to a genome editing tool, it has opened a new horizon in the field of biological sciences but it was a tough journey. One of the basic challenges faced by the scientists was to characterize various types of CRISPR Systems, their characterization was a big challenge thanks to extensive exchange of CAS genes and gene modules among various bacteria.

Scientists across the globe performed number of genomic analysis in order to establish a classification system capable of broadly identifying various types of CRISPR CAS systems. After that position specific scoring matrices (PSSM) of all the known CAS systems were developed. Then only those CAS locus were considered for further classification which were declared complete. A CRISPR-CAS system was declared as complete if it posses the adaptation modules and complete set of genes required for the formation of interference module. Keeping in view all these findings CRISPR-CAS system was divided on the basis of what type of interference module is encoded by its genes. Those encoding for a multi subunit CrRNA complex were placed in “Class-1” CRISPR systems and those encoding for a single multipurpose interfering module were placed in “Class-2” CRISPR-CAS systems. Each class has further types and subtypes.


The Class-1 has:  
  •  Type I
  •  Type III
  •  Type IV


The Class-2 has:  
  • Type II
  • Type V

Class 1:

The class 1 systems are described as those types of systems which require a large complex of multiple proteins to conduct the interference procedure. The class 1 has further 3 types The Type I, Type III and Type IV systems.
   v  Type I:
          The type I system is further divided in to a total of 7 subtypes namely sub-type, I-A, I-B, I-C, I-D, I-E, I-F, and I-U. All kinds of type I locus consists of a gene that encodes for the signature protein CAS 3 or its variant the CAS 3’. This furthers promotes the formation of helicase  which causes unwinding of double stranded DNA or the DNA-RNA complex. The helicase is fused with endonucleases which causes the cleavage of target DNA. Many of the subtypes are typically encoded by a single operon which encodes for cas1, cas2, cas3 and genes for cascade complex subunits.

  v  Type III:
             All the type III CAS systems are characterized by the presence of a specific protein called the CAS 10 protein which is the signature for the Type III systems. This CAS 10 induces formation of a multi-domain protein containing palm domain which is the largest subunit of the crRNA-effector complex. The type III systems are also reported to produce a CAS7 and a CAS5 protein subunit. This complex is further fused with specific nucleases which cuts and denatures the enzyme.

  v  Type IV:
              The type IV CAS system is one of the two new CRISPR CAS systems discovered in the recent studies. In this type the “csf1” protein serves as the signature protein. The specific function of this system type is still uncharacterized but how it functions? That has been sorted out recently; it has a multi-subunit crRNA-effector consisting of Csf1, Cas5 and Cas7 protein subunit. In addition to that it also contains a Ding family helicase or a alpha-helicase.

Class 2:

These type of systems are describes as those type of CRISPR-CAS system in which the entire procedure of interference is carried out by a single large proteinic molecule.  The class 2 has 2 types.
  v  Type II:
            Type II system is the most widely studied system because of the famous CAS9 belonging to this type. The signature gene for this system is the CAS9 which encodes for a single multi-domain protein which carries out the interference function. The system also contains CAS1 and CAS2 genes which carries out the adoption stage and is often assisted by the CAS9 protein. The type II system is further subdivided in to 3 subtypes; subtype II-A, II-B, and II-C. The II-A subtype posses an additional “csn2” gene, it has been reported to assist in spacer acquisition. The subtype II-B lacks csn2 but it has a “cas4” gene. The subtype II-C has only three genes the CAS1, CAS2 nad CAS9.
3D model of CAS9


  v  Type V:
            Type V is one of the two newly discovered CRISPR systems (type IV being the other newly discovered system). This system is characterized by having a special gene called the “cpf1” gene, which encodes for a large protein of about 1300 amino acids. This system type was first discovered in prevotella and francesella bacterial species. In similarity to CRISPR CAS9 all the interference process is conducted by a single large protein module named the CPF1. However the cpf1 lacks a n HNH nucleases domain which is common in CAS9 protein. Also the cpf1 is encoded outside the CRISPR-CAS context in several genomes which indicates it to be a possible addition because of transposable elements. In addition to cpf1 it also encodes for a cas1, cas2 and in rare cases cas4 proteins. Unlike other Class II systems which are specific to bacteria the cpf1 has been reported in one archae as well.
 
3D model of CPF1
Conclusion:
                         Till now scientists have successfully classified various CRISPR-CAS system in to 2 classes, 5 types and 16 subtypes, on the basis of their genetic characters and protein modules produced by the genes. This classification is vital for scientists as it will enable them to further enhance their knowledge, easily identifying and placing new CRISPR systems and implementing newly discovered systems for further high tech research.

CRISPR/CAS technology for making better sugarcane based biofuel products

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