MXPA00010830A - Method for prolonged storage of dna molecules and packaging implementing said method - Google Patents

Abstract

The invention concerns a method for prolonged storage of DNA molecules and a packaging for implementing said method. Said method is characterised in that, when the DNA has been extracted and purified by any suitable technique,whether standard or not, it consists in encapsulating the previously dehumidified DNA in a sealed, non-corrosive metal capsule (1). The invention is useful for storing DNA.

Description

PROCEDURE OF CONSERVATION OF LONG TERM OF MOLECULES OF DNA AND DEVICE TO MAKE IT FIELD AND ANTECEDENTS OF THE INVENTION The present invention refers to DNA (deoxyribonucleic acid), especially of human origin, and tending to preserve the DNA molecule which is the carrier of genes characteristic of each individual, that is, of their genetic patrimony. More precisely, the invention tends to preserve genetic information for the preservation of the DNA molecule as long as possible and under conditions of preservation of genetic information integrity. This information presents numerous interests especially for predictive medicine for genetic genealogy and identification. If the DNA molecule is relatively stable, the studies of archaeologists have revealed that it can be conserved millions of years in a favorable environment, however it can degrade in the absence of conservation in such an environment. Among the causes of DNA alteration can be mentioned the action of ionizing radiation, such as X-rays or gamma rays, the action of ultra violet radiation, oxidation and enzymatic or chemical hydrolysis.

SUMMARY OF THE INVENTION. The invention tends to propose a technique for preserving DNA in an environment, preserving it from the effects of the previous actions. To this end, the invention has as an object a long-term preservation method for DNA molecules, characterized in that it consists, after the extraction and purification of the DNA by any suitable conventional technique or not, of effecting an encapsulation in a sealed, stainless metal capsule. of the previously dehumed DNA molecule. DESCRIPTION OF THE INVENTION According to a first way of carrying out the process, the DNA is encapsulated in an atmosphere constituted by one or several inert gases and having a humidity degree less than or equal to 1 ppm of water. According to a second way of carrying out the process before the physical encapsulation, the DNA is subjected to a chemical encapsulation by wrapping it in an appropriate copolymer. According to a variant of this last embodiment, the chemical encapsulation is carried out with the aid of a hybrid consisting of said copolymer of organic molecules and / or inorganic salts for the purpose of reinforced protection of the DNA with respect to ultraviolet radiation or ionizing. Preferably, and whatever the method of carrying out the process, the physical encapsulation is completed by placing the sealed and stainless steel capsule in a container resistant to shocks and crushing. The DNA thus encapsulated can theoretically remain for several tens of thousands of years protected from the ultraviolet ionizing radiations of chemical aggressions and mechanical stresses. Another object of the invention is the different types of devices obtained according to the method, which are presented in the form of either a single capsule or a capsule enclosed in a protective envelope called a container. In the interior of the capsule, a quantity of DNA, for example 30 μ, is sufficient to carry out a substantial number of samples at all times. The DNA is deposited directly on the capsule or in a small glass container inserted into the capsule. In each sample, the capsule is opened, the desired amount of DNA is extracted, leaving the rest in the capsule, the hermetic device is reconstituted. The DNA removed below is rehydrated for analysis purposes. Such a device is of a nature that ensures a perennial preservation of the genetic heritage that is sheltered especially from oxidation and ionizing and ultraviolet radiation as well as from other chemical or mechanical aggressions. Other features and advantages will be seen from the description to be followed of the modes for carrying out the description procedure, description given only by way of example and with reference to the attached drawing, in which the single figure schematically illustrates a device structure according to the invention. Prior to carrying out the invention, the DNA is extracted and purified. This can be done by any conventional method or not. DNA can be extracted from any cell in the body. By way of example, DNA is extracted from blood, hair follicles, cellular follicles extracted from saliva, mucous membranes or skin cells. Next, a multi-stage purification process is carried out, namely: separation of the cells, elimination of proteins by enzymatic digestion, isolation / extraction of the DNA, amplification by polymerization, if necessary and conservation avoiding the factors of alteration. The DNA thus prepared is presented in the form of a precipitate in alcohol. The above protocol is well known and can be replaced by another existing or new procedure. The DNA according to the invention is then placed in a sealed, stainless metal capsule. Such a so-called physical encapsulation is carried out in an atmosphere formed of one or more inert gases such as rare gases and has a very low degree of humidity, preferably less than 1 ppm of water, said atmosphere being at atmospheric pressure. This is done, for example, with the aid in a conventional chamber or drying box. The capsule represented schematically in one on the figure of the attached drawing, is for example a gold capsule formed of a small circular bucket closed by a pressure-sensitive cap on the periphery of the bucket. The diameter of the gold capsule is, for example, of the order of 5 mm and its thickness of 2 to 3 mm. The hermetic unification of the tray and its lid can be carried out by any suitable means. Gold is preferred because of its malleability and its non-oxidation and non-DNA contamination properties. In a variant, an alloy based on gold or platinum can be used. The atmosphere of the encapsulation is dried in order to avoid hydrolysis and oxidation of the DNA after closure. The amount of DNA placed in the capsule, for example about thirty micrograms, is broad enough to allow a substantial number of different withdrawals of DNA from the capsule during its storage. Such a device may be sufficient to ensure the perennial preservation of the genetic heritage for tens and even hundreds of thousands of years, as long as it is understood that the capsule remains intact. This device protects the DNA in particular against: - chemical or enzymatic hydrolysis reactions; of disturbances induced by ultraviolet radiation or all ionizing radiation, such as X-rays or gamma rays; - of oxidation by oxygen from air. Advantageously and to reinforce the preservation of DNA the capsule l, that is the aforementioned gold capsule itself is enclosed in a sealed container 2, made of a material having good mechanical properties, in order to better protect the capsule 1 against mechanical aggressions, in particular the vibrations , the collisions and the crushing or any other aggression, for example a rise in temperature and in a general manner, so as to protect said capsule against the environment, whether normal or abnormal. The container or container 2 can be a kind of small box in two parts 2a, 2b, sealed or joined by any means to ensure the integrity and tightness of the assembly. The container 2 may for example be made of a suitable material, for example ceramic, a metal compound or a polymer. According to another way of carrying out the method of the invention, the DNA duly prepared before being put in place in the capsule 1, is subjected to a so-called chemical encapsulation. To this end, the DNA is wrapped in an inert polymer or copolymer in front of DNA, in order to reinforce the protection with respect to alteration factors, the DNA molecules are protected in the pores of the DNA. (co) polymer. The wrapping material is selected in such a way that it can be subsequently dissolved in order to be able to recover the DNA molecules.

All (co) polymer may be convenient, except those in which there is danger of the DNA molecule reacting those that would prevent the internal dissolution of DNA and those that would need for such redissolution an acid solvent, that is those that have a lower PH or equal to 4 approximately. Such chemical encapsulation can be performed by placing the DNA material properly prepared, for example a DNA ball obtained by precipitation in a solution of acrylic or polyacrylic acid in methyl alcohol. For example, a solution is placed in 50 cm3 of methyl alcohol, 1 g of acrylic or polyacrylic acid. The alcohol evaporates slowly until a viscous gel is obtained. A DNA accumulation of approximately 1 mm3 in suspension in an alcoholic solution is placed inside that gel. The whole is dried at 50 ° C until obtaining a solid block containing the DNA which is then placed in the capsule l according to the process described above, with the difference that it is no longer necessary to operate in a dry box, maintaining the dry atmosphere. Subsequently and at all times after the opening of the casings 1 and 2 the DNA can be de-encapsulated by immersion for one hour in ethyl alcohol, an accumulation of molecules identical to that obtained by the purification process is obtained again. According to an example horse using another (co) olimer, a solution of lg, methyl methacrylate or polymethacrylate dissolved in 50 cm3 of dichloromethane is prepared, the solvent is evaporated until obtaining a viscous gel, a DNA accumulation of approximately lmm3 in suspension in an alcoholic solution it is placed inside that gel. The whole is dried at 50 ° until obtaining a solid block containing the DNA which is placed in place in the capsule 1, under the same conditions as in the previous example. Subsequently and at all times after the opening of casings 1 and 2, the initial DNA can be regenerated by dissolving the copolymer (co) polymer by dichloromethane. According to a variant of such chemical encapsulations, the copolymer is associated with organic molecules and / or inorganic salts with the object of a better protection against ultra violet radiation and ionizing radiation, it is especially possible to add to the copolymer heavy metal ions in the measure in any case where there is no danger of DNA damage. According to one example, lg of acrylamide or polyacrylamide is dissolved in 25 cm3 of distilled water. 50mg of copper acetate and 50mg of zinc acetate are added to the solution. the water evaporates slowly until obtaining a viscous gel, a DNA accumulation of approximately lmm3 in suspension in an alcoholic solution is placed inside that gel. The whole is dried at 50 ° C, until obtaining a solid block containing the DNA, which is then placed in place in 1, under the same conditions as the preceding examples. It should be noted that the chemical encapsulation with the aid of a (co) polymer has the advantage of carrying out a polymerization of individual pellets, which makes subsequent DNA withdrawals more practical, since it is sufficient to remove a pellet or pellet from the capsule without touch the others. As the polymer which can be used according to the invention, it is also possible to mention agarose. Finally, the invention is obviously not limited to the illustrated embodiments but covers all variants, in particular as regards the materials constituting the capsules 1, or the containers or containers 2, the conditions of physical and chemical encapsulation. , the nature of the chemical encapsulation (co) polymer and its possible additives as well as the individual shapes of the capsules 1 or containers 2. Thus, in the variant, the DNA can be deposited on a glass layer, especially a soda-calcium glass , as schematized in 3, in Fig. 2, of the attached drawing. The DNA adheres to the glass of the small container 3, which can have, by way of example, a diameter of 7mm and a height of 1.2mm, the small container 3 being enclosed in a sealed manner in the capsule 1.

Claims (13)

1. NOVELTY OF THE INVENTION Having described the invention as above, the content of the following claims is claimed as property: CLAIMS 1.- Long-term preservation procedure of DNA molecules characterized in that it consists, after extraction and purification of the DNA, of any conventional technique or not, in effecting an encapsulation in a sealed, stainless metal capsule of the previously de-wetted DNA molecule.
2. 2. - Method according to claim 1, characterized in that the DNA is encapsulated in an atmosphere consisting of one or more inert gases and having a humidity level less than or equal to 1 ppm of water.
3. 3. Method according to claim 1, characterized in that before the physical encapsulation the DNA is subjected to a chemical encapsulation by wrapping in a suitable copolymer.
4. 4. Method according to claim 3, characterized in that the chemical encapsulation is carried out with the aid of a hybrid consisting of said copolymer of organic molecules and / or inorganic salts for the purpose of reinforced protection of the DNA with respect to the ultra violet or ionizing radiations.
5. 5. - Process according to claim 4, characterized in that the hybrid is composed of a copolymer and heavy metal ions.
6. 6. - Method according to one of claims 3 to 5, characterized in that the copolymer and its optional organic or inorganic additives are dissolved in a solvent evaporated afterwards until a viscous gel is obtained, then a determined set or accumulation of DNA in suspension in an alcoholic solution is placed inside the gel and the whole is dried at an appropriate temperature until obtaining a solid block containing the DNA.
7. 7. - Method according to claim 6, characterized in that the chemical encapsulation is followed by encapsulation in a hermetic, stainless metal capsule in an atmosphere formed of one or more inert gases.
8. 8. - Method according to one of claims 1 to 7, characterized in that the capsule is placed in a sealed container resistant to shock and crushing.
9. 9. - Method according to one of claims 1 to 8, characterized in that before its encapsulation in the metal capsule the DNA is deposited in a small glass container.
10. 10. Preservation device obtained according to the method according to one of claims 1 to 7, characterized in that it is constituted by a capsule of metal or malleable metal alloy and has non-oxidation and non-contamination properties of the DNA, closed in airtight manner by a cover by any suitable means, especially a boxing.
11. 11. - Device according to claim 10, more particularly intended to perform the method of claim 8, characterized in that it is enclosed in a container or container in a material selected from the group comprising ceramics, compounds, metals and polymers.
12. 12. - Device according to the claims 10, 11, more particularly intended to carry out the method of claim 9, characterized in that it has a small glass container on the inside of the metal capsule on which the DNA is deposited.
13. 13. - Device according to claim 12, characterized in that the small container is formed of a glass of calcium soda.