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Molecular marker method of two mutation sites in chicken FSHR (Follicle-Stimulating Hormone Receptor) gene 5' control region and applications thereof in children breeding

A technology of mutation sites and molecular markers, applied in the field of molecular genetics, can solve problems such as breeding and inability to apply chickens

Active Publication Date: 2011-06-01
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For poultry, especially the research on the 5' regulatory region of the chicken FSHR gene is still relatively small, and it cannot be applied to chicken breeding and other work.

Method used

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  • Molecular marker method of two mutation sites in chicken FSHR (Follicle-Stimulating Hormone Receptor) gene 5' control region and applications thereof in children breeding
  • Molecular marker method of two mutation sites in chicken FSHR (Follicle-Stimulating Hormone Receptor) gene 5' control region and applications thereof in children breeding
  • Molecular marker method of two mutation sites in chicken FSHR (Follicle-Stimulating Hormone Receptor) gene 5' control region and applications thereof in children breeding

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1 Cloning, Sequencing, Sequence Alignment and Mutation Site Analysis of Chicken FSHR 5' Regulatory Region Sequence

[0028] According to the published red jungle fowl sequence (GenBank Accession No.NW_060321), two sets of primers FRP1 (its sequence is shown in Seq ID No: 8 and Seq ID No: 9) and FRP2 (its sequence is shown in Seq ID No: 10 and Seq ID No: 10 and Seq ID No: 11), these two primers are specially designed according to the sequence of the red junglefowl registered in the database for the study of mutations in the chicken FSHR 5' regulatory region, and overlap 19bp to facilitate sequence assembly. Two pairs of primers respectively amplified four local breeds: Jining Hundred Day Chicken, Wenchang Chicken, Xianju Chicken, Tibetan Chicken and a commercial laying hen Hailan brown FSHR 5' regulatory region overlapping upstream (-1824bp~-940bp ) and downstream (-960bp ~ -40bp) two sequences. Fragment 1 amplified by the FRP1 primer is about 885bp ( figure 1 ...

Embodiment 2

[0035] Example 2 Polymorphisms of the two mutation sites -237 and -868 in the 5' regulatory region of chicken FSHR and their association with egg production traits

[0036] 1 test material

[0037] 36 Jining hundred-day chickens (Jining City, Shandong Province), 40 Wenchang chickens (Hainan Province), 29 Tibetan chickens (Tibet Autonomous Region) and 37 Roman browns (commercial chickens), 141 Lu birds (Shandong Zibo Mingfa breeders) field). There are 349 Wenchang chickens with egg production records (Hainan Luoniushan Wenchang Chicken Breeding Co., Ltd.), 390 new poplar brown 101 lines, and 204 new poplar brown heterozygous offspring (Shanghai Poultry Breeding Co., Ltd., National Poultry Engineering Technology Research Center) . All of the above were randomly sampled, blood was collected from the wing vein, anticoagulated with ACD, brought back at low temperature, and stored at -20°C.

[0038] 2 test method

[0039]2.1 Extraction of chicken blood DNA

[0040] Chicken bloo...

Embodiment 3

[0079] Example 3 Effect of Haplotype of Mutation Site in Chicken FSHR 5′ Regulatory Region on Regulated Gene Expression

[0080] 1 test material

[0081] Three sea blue and brown hens in the peak laying period of 29w were collected from the Animal Husbandry Experimental Station of Shandong Agricultural University for the cultivation of follicular granulosa cells.

[0082] 2 test method

[0083] 2.1 Construction of TG+ and TG- haplotype expression vectors

[0084]1) Select a new poplar brown hen individual whose genotype at the -237 site of the FSHR 5' regulatory region is TT, and the genotype at the -868 site is G+G- (that is, TTG+G-type), and its DNA is used as template to obtain TG+ and TG- haplotype sequences.

[0085] 2) Add the recognition sequence of the restriction endonuclease KpnI at the 5' end of the positive strand of the original P-868 primer, add the recognition sequence of the restriction endonuclease BglII at the 5' end of the negative strand, and add 3 -5 p...

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Abstract

The invention relates to the field of molecular genetics, in particular to a molecular marker method of two mutation sites in a chicken FSHR (Follicle-Stimulating Hormone Receptor) gene 5' control region and the applications thereof in children breeding. The inventor finds that chicken FSHR 5' control region -237 and -868 sites are not related to properties when single site marker is analyzed but finds that the two sites have remarkable difference (P=0.016) by corresponding a TTG+G- type to a small first laying age (123d) and corresponding a TTG+G+ type to a great first laying age (134d) when using the two site haplotypes to construct doubling types for multiple comparison. Accordingly, the invention can detect the two molecular markers related to the egg laying properties, and the method not only is simple, convenient and rapid, but also avoids environmental influence and can realize early seed section.

Description

technical field [0001] The invention relates to the field of molecular genetics, in particular to a molecular marker method for two mutation sites in the 5' regulatory region of chicken FSHR gene and its application in breeding. Background technique [0002] Follicle-stimulating hormone (FSH) is one of the important hormones that regulate the reproductive activities of animals. It plays an essential role in the growth, development, differentiation, maturation and ovulation of animal ovarian follicles. Its biological functions are performed through the FSH located on the target cell membrane. Receptor (FSHR) mediated. Studies have shown that the gene frequency of the polymorphic site at the 5′ end of the FSHR gene is significantly different in the Chinese Simmental cattle populations that gave birth to twins and single births. The study found that the level of FSHR mRNA in the developing follicle granulosa cells of multiple-born Romanov sheep was significantly higher than th...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 姜运良康丽张宁波唐辉张渝洁刘东君王慧梁森藏丽李标
Owner SHANDONG AGRICULTURAL UNIVERSITY
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