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Populus deltoids*populus nigra basic-domain leucine-zipper (PdbZIP) gene and application thereof

A gene, European and American technology, applied to the PdbZIP gene and its application field, to achieve the effect of improving expression

Inactive Publication Date: 2014-12-10
DALIAN NATIONALITIES UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, the bZIP gene of many plants has been extensively studied, but there is no report on the study of poplar in Europe and America

Method used

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  • Populus deltoids*populus nigra basic-domain leucine-zipper (PdbZIP) gene and application thereof
  • Populus deltoids*populus nigra basic-domain leucine-zipper (PdbZIP) gene and application thereof
  • Populus deltoids*populus nigra basic-domain leucine-zipper (PdbZIP) gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1 Extraction of total RNA

[0021] 1. The European and American Populus 107 material used in the present invention is taken from Baoding, Hebei. It is an annual cutting cuttage cultivated in the nursery of Beijing Forestry University. After the cuttage, it is watered once every three days in order to keep the soil moist. After new leaves grow, the growth similar (Growing for 3 months) European and American poplar seedlings were sprayed with 250mM NaCl, and the top leaves were removed at 0, 2, 4, and 6 hours respectively and placed in liquid nitrogen, and then stored in a -80°C ultra-low temperature refrigerator for later use. For total RNA extraction and cDNA synthesis.

[0022] 2. The process of extracting total RNA from the leaves of Populus americana under 250mM NaCl stress by CTAB method:

[0023] (1) Before extracting RNA, add DTT at a final concentration of 0.1 mM to 2×CTAB buffer, and preheat at 65°C.

[0024] (2) Put 0.5 g of liquid nitrogen ground mat...

Embodiment 2

[0030] Example 2 Using fluorescent quantitative PCR method to detect the expression of PdbZIP gene of Populus americana in adversity

[0031] According to the conserved sequence of PdbZIP gene of Populus americana, specific primers were designed: upstream primer: PdbZIP1 (SEQ ID NO.1) and downstream primer: PdbZIP2 (SEQ ID NO.2).

[0032] Primer name

sequence name

Base sequence (5'--3')

PdbZIP1

SEQ ID NO.1

ATGAGCCGCA TCTTCACAAC TCCTGAA

PdbZIP2

SEQ ID NO.2

TTAGCCTAGC AAATCTGAAG AACTTGTAAT

[0033]According to the method of Example 1, the cDNA samples prepared by extracting the apical leaves at 0, 2, 4, and 6 hours were used as templates, and PdbZIP1 (SEQ ID NO.1) and PdbZIP2 (SEQ ID NO.2) were used as templates. Primers were used for fluorescent quantitative PCR analysis.

[0034] The fluorescent quantitative PCR reaction system (20 μL):

[0035]

[0036] The fluorescent quantitative PCR reaction is divided into three s...

Embodiment 3

[0038] Example 3 Clone PdbZIP gene of Populus americana by PCR method

[0039] According to the conserved sequence of PdbZIP gene of Populus americana, specific primers were designed: upstream primer: PdbZIP1 (SEQ ID NO.1) and downstream primer: PdbZIP2 (SEQ ID NO.2).

[0040] Primer name

sequence name

Base sequence (5'--3')

PdbZIP1

SEQ ID NO.1

ATGAGCCGCA TCTTCACAAC TCCTGAA

PdbZIP2

SEQ ID NO.2

TTAGCCTAGC AAATCTGAAG AACTTGTAAT

[0041] The synthetic cDNA prepared by the method described in Example 1 was used as a template, and PdbZIP1 (SEQ ID NO.1) and PdbZIP2 (SEQ ID NO.2) were used as primers for PCR reaction;

[0042] The PCR reaction system is (25 μl):

[0043]

[0044] The PCR reaction program is as follows: 35 cycles of pre-denaturation at 95°C for 5 min, each cycle of denaturation at 95°C for 50 s, annealing at 63°C for 90 s, extension at 72°C for 2 min, and finally, the sample was extended at 72°C for 10 min. Th...

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Abstract

The invention discloses a stress related gene PdbZIP of populus deltoids*populus nigra (Populus deltoids*Populus nigra). The gene is very important in a salt-resistant process of a plant. The provided bZIP gene is named PdbZIP and has a base sequence shown in SEQ ID NO:3 in a sequence table. A promoter is added before initial nucleotide is transcribed when the stress related gene PdbZIP disclosed by the invention is built in an expression vector pCAMBIA1304; meanwhile, an optional marker green fluorescent protein (GFP) is added, so as to identify and screen transgenic plant cells or plants; the expression vector with the stress related gene PdbZIP disclosed by the invention can transform a plant host by many methods, and is used for cultivating salt-resistant plant variety. The gene disclosed by the invention has a wide application prospect in cultivation of the salt-resistant plant.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, in particular to a PdbZIP gene from Populus deltoides×Populus nigra and its application. Background technique [0002] European and American poplar (Populus deltoides×Populus nigra) is one of the most suitable tree species for short-rotation industrial timber intensive management in mid-latitude regions. In recent years, many excellent European and American poplar clones have been introduced into our country to create large-scale fast-growing and high-yielding forests and have achieved good economic and social benefits. But high salt etc. limit its further popularization. Therefore, when it is introduced into high-salt and water-scarce areas, screening and cultivating salt-resistant strains is a prerequisite. With the development of molecular biology, it has become an important way to solve this problem by using molecular biology techniques to study the molecular mechanism of salt r...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/11C12N15/82C12N1/21C07K14/415A01H5/00
Inventor 郭鹏董燕
Owner DALIAN NATIONALITIES UNIVERSITY
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