Highland bacillus altitudinis and application thereof in artificial ageing of tobacco leaves at upper part of flue-cured tobacco

A Bacillus, highland technology, applied in the field of tobacco aging and agricultural microorganisms, can solve the problems of poor quality and poor availability of tobacco leaves, and achieve the effects of reducing irritation and miscellaneous gas, improving aroma quality and aroma quantity, and shortening the aging period.

Inactive Publication Date: 2012-09-19
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to overcome the defects of the prior art, aim at the inferior quality and poor usability of the upper tobacco leaves of flue-cured tobacco, especially the aging of the upper tobacco leaves of flue-cured tobacco, and provide a new strain of microorganisms suitable for aging the upper tobacco leaves of flue-cured tobacco, namely Bacillus uplander , using this microorganism to improve the artificial aging of the upper tobacco leaves of flue-cured tobacco plants, so as to improve the quality of tobacco leaves

Method used

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  • Highland bacillus altitudinis and application thereof in artificial ageing of tobacco leaves at upper part of flue-cured tobacco
  • Highland bacillus altitudinis and application thereof in artificial ageing of tobacco leaves at upper part of flue-cured tobacco
  • Highland bacillus altitudinis and application thereof in artificial ageing of tobacco leaves at upper part of flue-cured tobacco

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1: Isolation and breeding of Bacillus highlander D2.

[0023] Tobacco leaves from different origins (Hubei Province, Yunnan Province) stored naturally for 2 years in the warehouse of Wuhan Cigarette Factory, and flue-cured tobacco leaves that had not been artificially aged were cut into pieces with sterilized scissors, soaked in 100 mL of sterile water, Shake on a shaker at 200rpm for 30min, take 0.1mL after shaking evenly, and spread it on an LB plate (medium composition (g / L): peptone 10g, yeast powder 5g, sodium chloride 10g, dilute to 1000mL with distilled water; agar 15-20g / L, adjust the pH of the medium to 7.0-7.4), after 12 hours of cultivation, number the various colonies, inoculate them in LB liquid medium, cultivate them for 12 hours, and inoculate them on the starch plate with an inoculation needle Above (medium composition (g / L): soluble starch 2g, peptone 10g, yeast powder 5g, sodium chloride 10g, distilled water to 1000mL, adjust the pH of the medi...

Embodiment 2

[0030] Example 2: Identification of the morphological and physiological and biochemical characteristics of the Bacillus alboteria (CCTCC NO: M2011056) D2 strain.

[0031] Refer to the test method of George M.Garrity "Bergey's Manual of Systematic Bacteriology" Vol.VIII (1974 edition) to detect its Gram staining, cell size and shape, presence or absence of flagella and spores, growth temperature, pH range, NaCl Tolerance, catalase, H2S test, starch hydrolysis, cellulose hydrolysis, MR test, phenylalanine deamination, urease test, oxidase, gelatin liquefaction, tyrosine degradation, V-P test, and utilization of mannitol, Arabica Sugar, galactose, xylose, rhamnose, sucrose, inositol and citrate tests.

[0032]The results showed that the bacterium was Gram-positive, rod-shaped, with a size of 0.5 μm to 1.0 μm × 3.0 μm to 6.0 μm, spores terminal or subterminal, perinatal flagella, and able to move; the growth temperature was 27-45 ° C, pH4~9, NaCl tolerance 2%~6%; gelatin liquefac...

Embodiment 3

[0033] Example 3: PCR amplification and sequence determination of the 16S rRNA gene of Bacillus highlander D2 strain.

[0034] The specific steps are:

[0035] Inoculate the D2 strain of Bacillus upaliensis (preservation number CCTCC NO: M2011056) in LB medium, 200rpm, 37°C shaking culture for 12 hours, centrifuge to collect the bacteria, add lysozyme after suspension, use CTAB method and SDS method to break the wall, use Genomic DNA was extracted with phenol-chloroform-isoamyl alcohol, and the 16S rRNA gene was detected with 16S rRNA universal primer forward primer 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and reverse primer 1492R (5'-GGTTACCTTGTTACGACTT-3'). PCR amplification, the amplified product was submitted to Shenzhen Huada Gene Research Institute for sequencing. The PCR conditions are: 94°C, 5min; 95°C, 40s, 55(-1°C / cycle)°C, 30s, 72°C, 25s, 6 cycles; 95°C, 40s, 55°C, 45s, 72°C, 1.5min, 30 cycles; 72°C, 5min, 10°C, 5min. The length of the PCR product is 1465bp, and its simi...

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Abstract

The invention belongs to the technical field of agricultural microbe and also relates to the technical field of tobacco leaf ageing. The invention discloses a microbial highland bacillus altitudinis D2 strain which is preserved in China Center for Type Culture Collection, the preserving number is CCTCCNO: m2011056, the invention further discloses the application of the highland bacillus altitudinis D2 strain in artificial ageing of tobacco leaves at the upper part of flue-cured tobacco. The highland bacillus altitudinis is fragtant adding and upgrading dominant bacteria separated from the surfaces of tobacco leaves, can ensure artificial ageing to tobacco leaves at the upper part of flue-cured tobaccom, shortens the aging time of the tobacco leaves at the upper part, improves the sucking quality of tobacco leaves at the upper part of flue-cured tobaccom, increases aroma quality and concentration of aroma, and reduces excitant.

Description

technical field [0001] The invention belongs to the technical field of agricultural microorganisms and the technical field of tobacco aging. Specifically, the invention relates to the use of an artificially isolated microbial strain, which is suitable for aging the upper tobacco leaves of flue-cured tobacco. The microorganism mentioned is Bacillus altitudinis D2. The invention also includes the application of the microorganism in the artificial aging of tobacco leaves on the upper part of flue-cured tobacco. Background technique [0002] Tobacco aging is an important process in the primary processing of tobacco, and aging is also an important link in the formation of tobacco smoking quality. Because the quality of the tobacco leaves (original tobacco) harvested in that year still has defects of varying degrees, such as the aroma is not prominent, the green smell is heavy, the irritation is strong, the taste is pungent, the aftertaste is not clean, the smoke is not delicate ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A24B15/20C12R1/07
Inventor 陈守文刘洋冀志霞韦杰李丹吴风光何结望李琳
Owner HUAZHONG AGRI UNIV
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