CN1306881A - Flowing separation process of horse and donkey X and Y sperm - Google Patents

Abstract

The present invention belongs to the sex control method in animal reproduction. X and Y sperms are separated in completely new method based on completely new theory. According to that X and Y sperms have different rotation and upstream property, diluted semen is filled into 100 cc injector and light irradiated to form closed convection, and in 40 min X and Y sperms are separated. It has a success rate of sex control over 90 %.

Description

Horse, donkey X, y sperm flow separation method

The present invention relates to horse, donkey X, y sperm flow separation method, belong to the sex-control method of animal reproduction aspect.

The data report, a nearly century has been studied to the sex control of domestic animal in countries in the world.Mainly be with the method differentiation X sperm of experiment and the delicate difference between y sperm, such as form, weight and the proportion of sperm, chemical analysis, to the reaction of soda acid, charged difference of institute and antigenicity etc., its method such as electrophoresis, density, acid-base method etc.But up to the present, also there is not a kind of effective method can make hundreds of millions of livestock semen samples be divided into x and y sperm for artificial insemination.

It is simple, easy to operate effectively to the purpose of this invention is to provide a kind of device, method, can realize the important breakthrough of modern genetics in theory and practice in the widely used horse of stud farm and breeding station, donkey X, y sperm flow separation method.

The sex of horse, donkey is the decision of do as one likes chromosome, is the male gamete abnormal shape.The quantity that contains the sperm generation of X, Y chromosome equates that therefore the probability that combines with ovum is also basic identical, thereby offspring's property ratio was near 1: 1.In view of the above, in the test-tube practice of horses, if the both sexes sperm of horse, donkey is separated, the just order ground do not controlled of attainability.

The sperm that utilization contains X, Y chromosome separately kinetic characteristic in fluid is separated it, according to having two:

Suitable, despining (spinning motion): according to observations and experiment, sperm is to rotate proally along its longitudinal axis, looks from afterbody, has clockwise and two kinds of inhours.The person of turning clockwise is a y sperm; The inhour revolver is the X sperm.

Sailing against the current property: in flow velocity was 1～2 times of sperm motility speed or slightly higher liquid stream, sperm had upstream characteristic.

Produce suitable, despun mechanism:

The normal sperm of domestic animal is to rotate proally along its longitudinal axis, and this just has report decades ago.But directionality problem about rotation, experiment (the 4th international livestock reproduction meeting translates 1961 selectively) according to Dutch E, M, BIOknuis, he thinks: " seeing that from behind up to the present the head of sperm does not see exception as yet with counter rotation (afterbody also rotates certainly) ".Repeatedly observe and test through the inventor, prove that this view only reflects the one side of problem, promptly except that counter rotation, clockwise in addition.

If the motion conditions that we observe horse, donkey, sperm with low power lens has four kinds of special phenomena just can find its movement locus:

One if a sperm is followed the tracks of in the mobile visual field, can find that then the movement locus of sperm is a garden shape, is not current generally acknowledged straight ahead motion.All sperms that have vigor normally all are (deformities, the bad ataxia person of causing of diluent preparing except) like this, and the rectilinear direction of its movement locus and motion (imaginary line) slightly departs from, and is arc-shaped bend.(dotted line is an imaginary line as Fig. 1 sperm arc track schematic diagram; Circle is represented the visual field).

Its two, the direction of the circus movement track of eupyrene sperm is divided into two kinds, a kind of is clockwise, a kind of is anticlockwise.

Its three, the ratio of the sperm count of clockwise and counter-clockwise circus movement track is 1: 1.

Its four, eupyrene sperm just moves to the direction of opposite circular arc after circular trace moves to a period of time, this is the result who touches slide.

Why does the movement locus of sperm have these phenomenons? observe according to the inventor, be to be caused by following three kinds of reasons: the profile of sperm is special, and sperm can rotate (rotation) motion and hydrodynamics.

1. the profile of sperm---separately, neck, tail three partly.Head is flat oval, intermediate recess; Neck is shorter, thin circular shaft shape; Afterbody is very long, is about 10 times of head length.It should be noted that between the swinging plane of the header planes of sperm of live body and afterbody an angle of cut is arranged.

2. rotation (rotation) motion

According to the observation, the forward travel of horse sperm is to lean on the rhythmic swing of afterbody to realize.Swing in wave shape, interact and reaction formation thrust with its surrounding liquid, when advancing, because the header planes of sperm and the swinging plane of afterbody have an angle of cut, and this angle of cut is suitable very cleverly, the liquid generation relative motion of the flat horizontal surface of head and side.Similar with kiteflying, produce pressure difference in face of the one side of liquid motion and opposing one side, and form lifting force.Under the effect of this lifting force, sperm head just raises up or following pressing down to the side.When head reaches new position, because afterbody and head are an integral body, also reach new position after being reversed thereupon, and continue swing.Follow this and go down, all constantly the shifting one's position in regular turn of space, sperm just rotates and has advanced for the swinging plane of afterbody and header planes.X, y sperm show that the relative position of tail swing plane and header planes is the axial symmetry that hand is levied formula, like this direction of rotation from afterbody look just have clockwise and anticlockwise different.

3. hydrodynamics

In static three-dimensional liquid, the movement locus of sperm is as follows:

When (1) sperm rotates forward travel, also be subjected to the influence of gravitational field, so its movement locus descends gradually.

(2) sperm owing to there are suitable, anti-two kinds to rotatablely move, under the situation of two kinds of sperm direction of advance unanimities, just following movement locus can occur in the process that rotation is advanced and descended;

First kind of medium---salt, carbohydrate

To contain y sperm is example, and the drop point that sperm descends is not in the projection of freely falling body, but skew left.This is because sperm is right handed, and the speed that rotate on its left side makes progress, and the speed that rotate on its right is downward, influenced in the process of decline by gravitational field, and the direction of the movement velocity of fluid (relative velocity) also is upwards.According to the speed addition principle, the left side has just become two speed sums like this, and the right has just become the poor of two speed.The pressure of " poor " be greater than " with " pressure, the movement locus that therefore contains the sperm of Y has just broken away from the track of freely falling body and skew left under the effect of this " additonal pressure ".In like manner, the movement locus that contains the sperm of X is offset to the right.

Second kind of medium---milk

Milk diluent is an emulsion with sugar, the salt dilution is different, abrim spherical particle.In the liquid of this irregular matter, the offset direction of sperm motility track is subjected to about two rightabout power institutes: the one, and " additonal pressure ", the one, " reaction force ".Along with the change of granular mass and concentration, it is identical with first kind of medium or on the contrary or three kinds of possibilities of non-migration that the offset direction of X, y sperm exists.

(3) in the process that sperm rotation is advanced, its direction of advance is not straight line but curve, and this is to descend owing to sperm is subjected to gravity, in the process that is subjected to " additonal pressure " and is offset, the shape unshapeliness of sperm itself, head is wideer several times than afterbody, and the suffered power of head is big.Under the effect of this kind vitality, the movement locus of its direction of advance is just crooked gradually, at last circle of Jin Si formation.Because suitable, anti-two kinds of rotation modes that sperm itself has, so circle also is suitable, anti-different commentaries on classics mark.

Sum up (1) (2) (3) each point, in static three-dimensional fluid, because the profile of X, y sperm structure has determined suitable, the despin campaign of sperm itself.Because suitable, anti-spinning motion has determined that again suitable, anti-two kinds of circular motion tracks and ratio thereof are 1: 1.

The upstream kinetic characteristic of sperm:

Normal sperm under suitable temperature, when running into the liquid stream of 1～2 times of own movement velocity all without exception the edge of a field sail against the current towards same direction.Flow stream velocity increases, the also corresponding quickening of the rotation number of sperm and movement velocity.

According to above-mentioned theory and practice, realize that the technical scheme of the object of the invention is:

1, with horse, donkey testicular fluid milk, sugared diluted;

2, the seminal fluid after will diluting extracts in syringe;

3, with syringe horizontal positioned light-illuminating;

4, X, y sperm close in syringe and are separated into two parts arrangement in the trivial circulation automatically, thus X, y sperm are separated.

The dilution prescription:

Sucrose 10%-12%

Milk powder 9%-11%

Accompanying drawing 1 is a sperm arc track schematic diagram, and dotted line is an imaginary line, and circle is represented the visual field.

Fig. 2 is X, y sperm flowing separating device structural representation.

Fig. 3 is the left view of Fig. 2.

Fig. 4 is that X, y sperm occupy the position schematic diagram.

Among the figure: 1-syringe, 2-separator frame, 3-light

Below in conjunction with accompanying drawing separation method of the present invention is further described. For realizing Separation method of the present invention is selected following equipment and power;

1. power---electric light light. This power more steadily, at the uniform velocity, be easy to control.

2. equipment---syringe, separator frame.

During separation syringe extracted that an amount of seminal fluid lies in a horizontal plane in the aluminum supporting spring On the vertical separation frame. At the certain distance on its opposite one light source being installed shines Penetrate, light and syringe are rectangular. Syringe is subjected to just luminous energy being turned to after the light irradiation Change into heat energy. The seminal fluid of dipped beam line is heated comparatively fast, its volume increase that density diminishes and on Rise, its position of vacating is filled by the cool bigger seminal fluid of density that stream comes. As This has alternately just produced convection current incessantly. Always there is temperature in the seminal fluid of dipped beam line and distance light line Difference, so convection current just continues not stop. This to flow phenomenon in the postradiation several seconds of light Just can start within the kind.

Syringe needs horizontal positioned, and the direction of liquid flow could be ring with ground like this Shape is vertical. Because syringe is filled a certain amount of seminal fluid, do not interspace, so seminal fluid In syringe, close trivial circulating take tube wall as outer Jie as circle.

Seminal fluid is viscous liquid, has internal friction, so outer field flowing must be drawn Playing internal layer flows accordingly. The flow velocity of seminal fluid is very little, and its inner liquid stream is the laminar flow shape Attitude. It is zero that outermost layer flow velocity maximum, middle shaft part are divided flow velocity, and there is a speed ladder centre Degree. According to mechanics principle, flow rate of liquid greatly then momentum is big; The little then momentum of flow velocity also Little. Sperm at the volley head the place ahead forms toroidal, in upstream process In, the effect that horn mouth is subjected to the difference of liquid amount of flow is flushed to slower one deck gradually. Follow this and go down, sperm is just passive to be moved to central axis direction. Meanwhile, because smart The rotation of son and produce " additonal pressure " because suitable, anti-clockwise difference and Separate to the syringe two ends.

Separation method:

Separation should be carried out in 18-25 ℃ darkroom.

At first make semen diluent:

With 11% sucrose 50ml and 10% milk powder 50ml mixed in equal amounts, put into same three In the flask of angle, moise-heat sterilization 10 minutes, take out be cooled to 37-40 ℃ for subsequent use. Separator has a separator frame, is the vertical plank of usheing to seat. Lay three rows on the plank The supporting spring of aluminum, it highly is respectively 7,13,19cm, puts syringe and uses. To identity distance 43cm place, put the bulb of a 300W, apart from height of table top at separator frame 5.6cm.

After taking horse or donkey testicular fluid and having measured semen volume, use immediately dilution with Dilution in 1: 1. With the seminal fluid 30ml after the 100ml syringe extraction dilution; In the seminal fluid Stay minute bubbles (so that water intaking is flat, being equivalent to the bubble in the level meter), its diameter 3mm is advisable. Syringe port stops up with the splint of having sterilized. This syringe is placed On the supporting spring of separator frame. Regulate the bubble in the syringe, make it to be in liquid level in The centre. Syringe is vertical with light. Turn on the power switch, shone 40 minutes. So After splint is pulled out gently, annotate keeping syringe to push in the situation stably as far as possible Penetrate the device handle, first half be collected in the seminal fluid bottle, surplus Lower Half be collected in another with In the seminal fluid bottle of mark. What be separated occupies the position of syringe with X, Y seminal fluid See Fig. 4: X, y sperm occupy the position schematic diagram.

According to the breeding needs, the seminal fluid after separating diluted again once carry out semen deposition.

The effect of invention sees Table 1.

The inventor from 1976 to nineteen eighty-two inner maintaining secrecy implement the effect that X, y sperm are separated of separation method success of the present invention, its sex is controlled to power up to 90.9%

Table 1: horse, donkey sex control result

Year	The foaling number	The sperm classification	The female-male proportion of foaling	The property control success percentage
					1976	9	X	7∶2	78.0
1977	29	X	21∶8	72.4
					1978	37	X	26∶11	70.2
1979	21	Y	16∶5	76.2
					1982	44	X	40∶4	90.9

Embodiment:

This invention should be carried out in kind of a horse (donkey) field or breeding station.The work of should getting ready earlier before X, y sperm separate.Be under 18-25 ℃ the condition in room temperature, mainly be 1, prepare dilution: 50 milliliters of 11% sucrose; 10% milk powder

50 milliliters; The two is 100 milliliters after mixing.And be adjusted at 37 ℃-40 ℃.

2, place separator, make separator frame and 300W light source keep 43 centimetres.5.6 centimetres of light height.3, will adopt seminal fluid with dilution by dilution in 1: 1, extract 30 milliliters of rare back seminal fluid with the 100cc syringe afterwards.Stay a small bubble in the seminal fluid, it directly is advisable through 3mm.The splint of syringe port after with sterilization stops up, be placed on the separator frame of band aluminium flake holder (divide three lattice on the separator frame: 7,13,19 centimetres, every lattice all can), regulate the vertical also maintenance level of syringe with light.Photoswitch turns on the light.After shining 40 fens kinds, splint is pulled out gently.Touch the syringe handle, with the first half seminal fluid collecting in the seminal fluid bottle that indicates Y, latter half of being collected in another seminal fluid bottle that indicates X.According to the needs of breeding quantity, the seminal fluid after separating is diluted once again, and choose required X or the Y seminal fluid of sex control mare is carried out semen deposition.The artificial insemination program is omitted.

Claims (3)

1, horse, donkey X, y sperm flow separation method utilize X, y sperm that suitable, despun difference and sailing against the current property are arranged, and it is characterized in that separation method is:

(1) with horse, donkey testicular fluid milk, sugared diluted;

(2) seminal fluid after will diluting extracts in syringe;

(3) with syringe horizontal positioned light-illuminating;

(4) X, y sperm close in syringe and are separated into two parts arrangement in the trivial circulation automatically, thus X, y sperm are separated.

2, horse as claimed in claim 1, donkey X, y sperm flow separation method is characterized in that the dilution prescription is: sucrose 10%-12%

Milk powder 9%-11%

3,, it is characterized in that separating according to following method according to horse, donkey X, the y sperm flow separation method of claim 1:

(1) with horse, donkey testicular fluid with 50 milliliters of 11% sucrose, 50 milliliters of dilutions of 10% milk powder; By dilution in 1: 1;

(2) with the seminal fluid after 30 milliliters of dilutions of 100 milliliters of syringes extractions;

(3) with the syringe horizontal positioned, with 300W electric light light, apart from syringe 43 cm distance to the syringe vertical irradiation;

(4) irradiation is after 40 minutes, and X, y sperm separate automatically.

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