CN105176802A - Bacterial colony density adjustable high-precision inoculation and photoprint tool - Google Patents

Abstract

The invention belongs to the microbial technology field, and concretely relates to a bacterial colony density adjustable high-precision inoculation and photoprint tool. The tool comprises a base plate, a middle plate and a face plate, and the base plate, the middle plate and the face plate are disc-shaped and have the same size. The base plate, the middle plate and the face plate are fixed through four stainless steel screws uniformly distributed at the edge, cooper studs sleeve the four stainless steel screws respectively. The centers of the base plate, the middle plate and the face plate are fixedly connected with a handle through nuts. 1448 micropores are arranged in the middle plate and the face plate, 10-1448 flat stainless steel nails go through the micropores in the middle plate and the face plate, and the nail heads can move up and down between the base plate and the middle plate. Through the tool, bacterial colonies can be inoculated in a quantificational manner, bacterial colonies can be photoprinted and copied with high precision, and firing disinfection can be carried out rapidly.

Description

High-precision inoculation and photocopying tool with adjustable colony density

technical field

The invention belongs to the technical field of microorganisms, and in particular relates to a high-precision inoculation and photocopying tool with adjustable colony density.

Background technique

In 1952, J. Lederberg invented the Replicaplating method, that is, through the inoculation and culture method of the replica plate, colonies of the same genetic type are formed on the corresponding positions of the mother and daughter plates. The lithographic method not only has an important application in the theoretical research of microbial genetics, but also has applications in breeding practice and other research. The traditional photocopying method is to stretch sterilized velvet on a cylinder slightly smaller than the petri dish to form a stamp-like inoculation tool, and then turn the petri dish with colonies upside down on the velvet stamp and print it lightly. An inoculation method in which colonies are transferred to velvet and transferred to another Petri dish containing medium by pressing. The advantage of the lithography method is that it facilitates large-scale screening of bacteria.

The shortcomings of the current photocopying tools are mainly as follows: 1. No inoculation function: the current photocopying tools cannot control the density of inoculated microorganisms, the spacing cannot be adjusted, and the growth speed of microorganisms is inconsistent, and the expansion speed of colonies is also inconsistent, resulting in the inability to be used for initial inoculation. . 2. Low photocopying accuracy: when using a stamp tool for photocopying, colonies at close distances are easily squeezed, causing contamination between colonies; when using a toothpick tool for photocopying, if there are colonies between the toothpicks, they cannot be photocopied, resulting in missing prints; use toothpicks When the tool is photocopied, a large colony may be multiplied into multiple colonies; when the surface of the plate medium is uneven, the stamp/toothpick photocopying tool will cause missing prints/puncture the medium respectively. 3. Long sterilization time: Since flannelette, toothpicks, bristles, etc. cannot be sterilized by burning on the flame, it is necessary to sterilize a large number of photocopying tools in advance during batch operation (each tool must be sterilized by damp heat again after photocopying once). Bacteria can be used for another motherboard), and a single moist heat sterilization takes 1 to 2 hours.

Contents of the invention

The technical problem to be solved by the present invention is to provide a high-precision inoculation and photocopying tool with adjustable colony density. Using this tool, colonies can be inoculated quantitatively, colonies can be reproduced with high precision, and it can be quickly sterilized by burning.

The inoculation and photocopying tool of the present invention comprises disc-shaped, same-size base plate, middle plate and panel, they are fixed by four stainless steel screws evenly distributed on the edge, copper studs are covered in four stainless steel screws, base plate, middle plate 1. The center of the panel and a handle are fixedly connected by nuts; there are 1448 small holes on the middle plate and the front panel, and 10~1448 flat-headed stainless steel nails pass through the small holes of the middle plate and the front panel, and the nail heads are on the bottom plate and the middle plate can move up and down.

The distance between the middle plate and the bottom plate is 4mm.

The distance between the middle plate and the panel is 8mm.

The bottom plate, middle plate, and panel are 1.6mm thick and 95mm in diameter.

Flat head stainless steel nails are 30±1mm long and 1mm in diameter. 10~1448 flat-headed stainless steel nails, which can be freely adjusted according to the size of the microbial colony. In order to avoid the mutual adhesion and fusion of the colonies while taking into account the separation efficiency, the number of needles should be less when the colony is large, and the number of needles should be more when the colony is small.

The function of quantitatively inoculating bacterial colonies of the device of the present invention refers to: according to the difference of the size of bacterial colonies, the maximum number of colonies that can be formed on each petri dish with a diameter of 10 cm is continuously adjustable between 10 and 1448; the function of high-precision photocopying of bacterial colonies refers to : The colony formed by inoculation of the same tool can be effectively copied to another plate, avoiding missing printing and printing, and the photocopying part (steel needle) is completely attached to the surface of the plate; Sterilization performance refers to: the whole tool can withstand 121 ℃ humid heat Sterilization, the part of the inoculation needle can be directly sterilized by rapid burning with the flame of an alcohol lamp.

The comparison between the tool of the present invention and the existing tool is summarized in the table above.

Description of drawings

Figure 1, a schematic diagram of the structure of the panel and the middle panel.

Fig. 2 is a schematic diagram of the structure of the device of the present invention.

Figure 3. Case of microorganisms in sediment, primary plate, showing the effect of inoculation.

Fig. 4, case of microorganisms in sediment, progeny plate, showing photocopying effect.

Figure 5, the case of ultraviolet mutagenesis of actinomycetes, the primary plate, showing the effect of inoculation.

Figure 6, a case of ultraviolet mutagenesis of actinomycetes, offspring plate, showing photocopy effect.

Detailed ways

One, the preparation of tool of the present invention

The panel and middle panel design are shown in 1. Among them, 1. Disk: diameter 95mm, thickness 1.6mm

2. Center hole: a small hole with a diameter of 3mm, located in the center of the disc, a total of 1;

3. Central axis holes: small holes with a diameter of 2 mm, located on the vertical and horizontal central axis of the yellow disc, with the center of the circle 5 mm from the edge of the disc, 4 in total;

4. Small holes: Drill small holes with a diameter of 1.15mm on the disc, and the distance between the vertical and horizontal holes (the distance between the centers of the circles) is 2.15mm, a total of 1448;

5. Positioning hole: a small hole with a diameter of 1 mm, located on the transverse central axis of the disc, and the center of the circle is 1.5 mm from the border of the disc, 1 in total.

There are 1448 small holes in the bottom plate, the diameter of the central hole is increased to 7mm, and the diameter of the central axis is increased to 4mm, and the others are the same as the panel.

The number of pins can be continuously adjusted between 10 and 1448 holes. The installation process takes 357 pins as an example: the two boards should be separated by copper studs with a distance of 8mm. Insert 357 flat-head stainless steel nails into the middle board and the front panel. Since the diameter of the stainless steel nail is slightly smaller than the diameter of the hole, the nail can move up and down in the hole; the bottom plate is assembled after the stainless steel nail is installed, and the function of the bottom plate is to limit the movement range of the stainless steel nail and force all the stainless steel nails to slide down naturally through its own gravity ( In order to ensure that the stainless steel nails and the plate fit well), the bottom plate and the middle plate are separated by copper studs, and the distance between the bottom plate and the middle panel is 0~4mm; then the bottom plate, the middle plate, and the panel are fixed with stainless steel screws, and then the handle is installed . The whole device can withstand 121°C damp heat sterilization for 1,000 times, and the inoculation site (nail tip) can be sterilized by burning with an alcohol lamp.

Two, two examples of the application of the tool of the present invention

Example 1. Isolation and photocopying of microorganisms in sediment

1. Sample source and culture medium

The source of the sample for the isolated microorganisms: the sediment from a reservoir in Hubei Province.

Medium: soluble starch 20g, KNO ₃ 1g, NaCl 0.5g, K ₂ HPO ₄ 0.5g, MgSO ₄ 0.5g, FeSO ₄ 0.01g, agar 20g, distilled water 1L, pH=7.2, agar powder content 1.6%.

2. Preparation of bottom sludge suspension

Weigh 1g of bottom mud into the Erlenmeyer flask, then add 50ml of sterilized physiological saline, then add glass beads, place in a shaker, shake at 25°C and 160r/min for 1h, and then let it stand for later use.

3. Plate photocopying inoculation bacteria (preparation of primary plates)

Take a 10cm petri dish sterilized at 121°C and put it in a clean workbench for 30 minutes of ultraviolet irradiation, take >10mL of the bottom sludge bacterial suspension and pour it into a clean sterilized petri dish, dip the bacterial suspension with an inoculation photocopying tool Finally, place it (depending on the tool's own gravity, no need to press) on the pre-prepared Gaoshi No. 1 solid plate (10cm petri dish), mark the positioning hole on the edge of the petri dish corresponding to the positioning hole of the inoculation photocopying tool with a marker pen, cover Incubate upside down at 25°C with the lid on the plate.

4. Plate photocopying and subculture (preparation of offspring plates)

When the colony on the plate grows to an appropriate size (the colony is obvious but not yet cohesive and fused), use aseptic operation, align the positioning hole of the same inoculation photocopying tool with the positioning hole mark on the petri dish, and put down the inoculation photocopying tool (depending on the tool’s Its own gravity, no need to press), the steel needle will be stained with the corresponding colony on the original plate, and then transferred to another Petri dish containing Gao’s No. Mark the positioning hole), cover the plate and culture it upside down at 25°C. After photocopying, the primary plates were stored at 4°C until use.

Example 2. Isolation and photocopying of actinomycete mutation breeding

1. Sample source and culture medium

The sample source of isolated microorganisms: a pure strain of actinomycetes -- Streptomyces AN02.

Medium: soluble starch 20g, KNO ₃ 1g, NaCl 0.5g, K ₂ HPO ₄ 0.5g, MgSO ₄ 0.5g, FeSO ₄ 0.01g, agar 20g, distilled water 1L, pH=7.2, agar powder content 1.6%.

2. Ultraviolet mutagenesis treatment of bacterial suspension

Place the bacterial suspension at a distance of 25 cm from the PHILIPSPL-S9W/01/2P medium-wave ultraviolet lamp for 3 minutes.

3. Plate photocopying inoculation bacteria (preparation of primary plates)

Take a 10cm petri dish sterilized at 121°C and put it in a clean workbench for 30 minutes of ultraviolet irradiation, take >10mL of the bottom sludge suspension and pour it into a clean sterilized petri dish, and use an inoculation photocopying tool to dip in the ultraviolet mutagenic Place the treated bacterial suspension (depending on the tool's own gravity, no need to press) on the pre-prepared Goose No. 1 solid plate (10cm petri dish), and use a marker pen to mark the edge of the petri dish corresponding to the positioning hole of the inoculation photocopying tool. Position the well marks, cover the plate and incubate at 25°C upside down.

4. Plate photocopying and subculture (preparation of offspring plates)

When the colony on the plate grows to an appropriate size (the colony is obvious but not yet cohesive and fused), use aseptic operation, align the positioning hole of the same inoculation photocopying tool with the positioning hole mark on the petri dish, and put down the inoculation photocopying tool (depending on the tool’s Its own gravity, no need to press), the steel needle will be stained with the corresponding colony on the original plate, and then transferred to another Petri dish containing Gao’s No. Mark the positioning hole), cover the plate and culture it upside down at 25°C. After photocopying, the primary plates were stored at 4°C until use.

Claims (5)

1. A high-precision inoculation and photocopying tool with adjustable colony density is characterized in that it comprises disc-shaped, same-sized base plates, middle plates and panels, which are fixed by four stainless steel screws evenly distributed on the edges, A stainless steel screw is covered with a copper stud, and the center of the bottom plate, middle plate, and panel is fixedly connected with a handle through a nut; there are 1448 small holes on the middle plate and the panel, and 10~1448 flat-head stainless steel nails pass through the middle plate and the panel The small hole, and the nail head can move up and down between the bottom plate and the middle plate. 2. The high-precision inoculation and photocopying tool according to claim 1, characterized in that the bottom plate is movable and presses all the stainless steel nails to slide down naturally through its own gravity (so as to ensure a good fit between the stainless steel nails and the plate). 3. The high-precision inoculation and photocopying tool according to claim 1, characterized in that the stainless steel nail is a flat head, its diameter is slightly smaller than the diameter of the hole, and the nail can move up and down in a small range in the hole. 4. The high-precision inoculation and photocopying tool according to claim 1, characterized in that the number of flat-headed stainless steel nails and the gap between the needles that can be equipped with the bottom plate and the middle plate can be adjusted freely according to needs. 5. The high-precision inoculation and photocopying tool according to claim 1, characterized in that, the tool as a whole can withstand damp heat sterilization, and the stainless steel nail can withstand alcohol lamp flame burning sterilization; In addition, the overall dimensions of the middle plate, front panel, and bottom plate designed by this tool, the aperture size and density of the holes, and the material, diameter, and length of the stainless steel nails can be adjusted according to needs; For example, a large petri dish with a diameter of 20 cm can be used when separating molds (very large colonies). The density decreases, the length of the stainless steel nail increases, and the material of the stainless steel needle can also be replaced by heat-resistant stainless materials such as aluminum needles and copper needles.