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Primer, kit and method for identifying dove sex

A kit, pigeon technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., can solve problems such as insufficient specificity and false negatives, and achieve good specificity, high recognition, high resolution effect

Active Publication Date: 2016-09-07
深圳市天翔达鸽业有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Reduce the detection cost, increase the success rate, lower the technical threshold, and bring revolutionary changes to the production of pigeons and even the production of poultry; there are some primers and methods for identifying the sex of pigeons in the prior art, but some of the disclosed primers are only amplified One band is used to identify the sex of pigeons by different band sizes, which has insufficient specificity and even the possibility of false negatives

Method used

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  • Primer, kit and method for identifying dove sex
  • Primer, kit and method for identifying dove sex
  • Primer, kit and method for identifying dove sex

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Primer design and specificity experiments

[0042]Venous blood sample processing method: Take 5 microliters of blood with a pipette in the early stage, and dilute it into tube A containing blood sample lysate according to the volume ratio of 1:30.

[0043] Blood sample processing method in feather follicles: Press the newly picked feathers along the feathers to the hair bulb to collect 10 microliters or more of fresh blood, and add 150 microliters of blood sample lysate.

[0044] Feather sample processing method: cut the collected feathers along the root to 3-4 mm, place in tube B containing 150 microliters of feather sample lysate, and place at room temperature for 2 minutes.

[0045] The PCR amplification procedure is as follows:

[0046] (1) According to the preset program in the PCR machine, heat at 98°C for 10 minutes, then cool at 4°C for ten minutes, take out tube A or tube B, 1.2×10 4 centrifuge for 10 minutes.

[0047] (2) Transfer the supernatant...

Embodiment 2

[0055] Example 2 Lysate Screening Experiment (Blood Sample)

[0056] The sample processing method is the same as in Example 1 (blood sample is selected here), and the PCR amplification procedure is as follows:

[0057] (1) According to the preset program in the PCR machine, heat at 98°C for 10 minutes, then cool at 4°C for ten minutes, take out tube A or tube B, 1.2×10 4 centrifuge for 10 minutes.

[0058] (2) Transfer the supernatant in tube A or tube B to a new PCR tube, take 2 microliters as a template for the PCR reaction, and then add 10 microliters of PermixTaq in sequence TM， , 6 microliters of double-distilled water, and 1 microliter each of the upstream and downstream primers, totaling 20 microliters of the system (the sequences of the upstream and downstream primers are shown in SEQ ID NO: 1 and SEQ ID NO: 2).

[0059] SEQ ID NO: 1: 5'-TTCTGAGGATGGAAATGAGT-3'

[0060] SEQ ID NO: 2: 5'-AGCAATGGTTACAACACTTC-3'

[0061] (3) Put the added system into the PCR machine...

Embodiment 3

[0067] Example 3 Lysate screening experiment (feather sample)

[0068] Pigeon feathers are rich in protein components, which brings great difficulty to effective amplification; we have explored a variety of feather-like lysates in the early stage, among which PBS buffer is very feasible as a feather-like lysate, The result of its amplification is as Figure 9 .

[0069] Next, the concentration of the experimental PBS buffer is the effect of the amplification result, the result is as follows Figure 10 , and finally determine 1×PBS as the lysate of pigeon feather samples. Using 1×PBS as the lysate, SEQ ID NO: 1 and SEQ ID NO: 2 as primers, verified with 30 male and female pigeons respectively, the results are as follows Figure 11 with 12 .

[0070] Depend on Figure 11 with Figure 12 We can know that after the feather samples are treated with 1×PBS, the PCR reaction and agarose gel electrophoresis can amplify the bands for distinguishing sex. but, Figure 12 The elec...

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Abstract

The invention discloses a primer, a kit and a method for identifying dove sex. The primer has sequences as shown in SEQ ID NO: 1 and SEQ ID NO: 2; the primer has the advantages of being good in specificity, high in resolution ratio, good in amplification effect and the like; and the primer, without enzyme digestion, can directly conduct agarose gel electrophoresis. Moreover, two specific products, compared with a specific product, are higher in recognition degree, so that an amplification result can be recognized at a glance; the invention also provides a method for identifying the dove sex by virtue of the primer; the method, under the precondition of conducting specific amplification by virtue of the primer, can also reduce time cost and economic cost of identifying the dove sex by simplifying a sample DNA acquisition process; and the method, which identifies the dove sex by virtue of the primer, is broad in market utilization value.

Description

technical field [0001] The invention relates to the technical field of molecular biology detection, in particular to a primer, a kit and an identification method for identifying the sex of pigeons. Background technique [0002] Gender identification is particularly important in poultry breeding, control of breeding costs and other production links. Individuals of different genders have different nutritional needs, and early gender distinction can be used to raise individuals differently; for breeding pigeons, a reasonable sex ratio is required, and early sex identification will help to achieve this process. [0003] Pigeons are monomorphic birds whose sexes are difficult to distinguish in appearance and other behaviors, whether they are juveniles or adults. The anus-turning identification method is currently the most widely used method in poultry production, but the newborn pigeons are small in size, only about 10g, and it is difficult to conduct sex identification in the e...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6879
Inventor 王修启梁少杰陈明霞高春起严会超
Owner 深圳市天翔达鸽业有限公司
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