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Specific molecular marker capable of identifying sea-tangle female and male gametophytes

A male gametophyte and molecular marker technology, applied in the field of genetic engineering

Active Publication Date: 2010-06-09
SHANGHAI OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no report on the specific molecular markers of the female and male daughters of Laminaria at home and abroad.

Method used

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  • Specific molecular marker capable of identifying sea-tangle female and male gametophytes
  • Specific molecular marker capable of identifying sea-tangle female and male gametophytes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0016] 1) Take the clonal line of Laminaria gametophyte with a fresh weight of 0.1 g.

[0017] 2) Gently grind the gametophytic clones into powder in liquid nitrogen, and immediately transfer to preheated 0.6ml CTAB extraction buffer containing 3% CTAB, 1.4M NaCl, 20mM EDTA in a weight-volume ratio , 10 mM Tris-HCl with a pH of 7.8 and mercaptoethanol with a weight to volume ratio of 2%, and mixed on a vortex mixer for 1 min.

[0018] 3) Warm bath in a 65°C water bath for 30-60 minutes, and gently shake it upside down every 5 minutes.

[0019] 4) Add an equal volume of chloroform: foreign alcohol mixture with a volume ratio of 24:1, and gently invert and mix for 3-5 minutes until an emulsified layer appears.

[0020] 5) Centrifuge at 12 000 rpm (rotation / minute) for 10 min at room temperature, and transfer the supernatant to another sterilized centrifuge tube.

[0021] 6) Add 2 / 3 volume of isopropanol pre-cooled at -20°C to the centrifuge tube, invert and mix until precipita...

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Abstract

The invention belongs to the technology field of the gene engineering, in particular relates to a specificity molecule marker which can distinguish laminaria female and male gametophyte. The specificity molecule marker is obtained through the following method: two pairs of primers of SEQ ID No1and SEQ ID No2 are designed according to 5'end flanking sequence of the laminaria female and female gametophyte gene groups, polymerase chain reaction amplification is performed to the deoxyribonucleic acid of the laminaria female and female gametophyte gene groups, and the deoxyribonucleic acid of specificity amplification fragment are reclaimed; large intestine rod competent strain JM109 is cloned through pMD19-T vector to obtain a laminaria female gametophyte specificity molecule marker Rf-763 anda laminaria male gametophyte specificity molecule marker Rm-239, and the sequences thereof are SEQ ID No3 and SEQ ID No4. The genders of the laminaria gametophyte, parthenogenesis laminaria sporophyte parental source and the genders of the offspring thereof can identified through utilizing the specificity molecule marker, and the laminaria gender differentiation in basis of the cell and moleculelevel can be studied.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and relates to a specific marker for determining the sex of a plant, in particular to a specific molecular marker capable of distinguishing the female and male gametophytes of Laminaria, using these molecular markers, the sex of the gametophytes of Laminaria can be quickly and effectively identified, and also It can be used to identify the source of the parental source of parthenogenetic Laminaria sporophytes and the sex of their offspring. Background technique [0002] For most plants, gender identification is mainly carried out through the external morphology of the plant, physiological and biochemical differences, chemical agent treatment, isozyme map, specific protein, karyotype, nucleic acid and other aspects. With the development of molecular biotechnology, especially the maturity of molecular marker technology, the role of molecular markers in plant sex identification is becomi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/29C12N15/11C12N15/10
Inventor 周志刚刘艳省毕燕会李利华
Owner SHANGHAI OCEAN UNIV
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