CN106906165B - Lactobacillus plantarum and application thereof in preparation of food for preventing constipation - Google Patents
Lactobacillus plantarum and application thereof in preparation of food for preventing constipation Download PDFInfo
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- CN106906165B CN106906165B CN201710278401.7A CN201710278401A CN106906165B CN 106906165 B CN106906165 B CN 106906165B CN 201710278401 A CN201710278401 A CN 201710278401A CN 106906165 B CN106906165 B CN 106906165B
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- lactobacillus plantarum
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- lactobacillus
- plantarum
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Abstract
The invention discloses Lactobacillus plantarum YS1 (L actinobacillus plantarum YS1) with the preservation number of CCTCC NO: M2016747 and application thereof in preparing food for preventing constipation, which not only expands the application range of the Lactobacillus plantarum YS1 and improves the application value thereof, but also brings a new hope for preventing constipation.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and relates to lactobacillus plantarum and application thereof in preparation of food.
Background
The yak yogurt is a natural fermented dairy product in Tibetan regions of Qinghai-Tibet plateau, contains rich nutrient components, and has the functions of resisting oxidation, reducing cholesterol and regulating immunity. The yak yogurt has special flavor and quality due to special natural fermentation conditions including raw milk fermentation, fermentation temperature, fermentation time and fermentation vessel, in particular to special fermentation microorganism composition.
When the frequency of defecation is less than 3 times per week, the constipation is considered to be in a constipation state, and when the frequency is less than 1 time, the constipation is considered to be a serious constipation state, which is a complex symptom threatening the health of human bodies, particularly having great influence on the health of colon. Constipation is not generally considered a disease, and most cases can be prevented by regulation of diet and habits.
Disclosure of Invention
The invention aims to separate and identify microorganisms in yak yogurt and research the activity and application of the microorganisms.
Through research, the invention provides the following technical scheme:
1. lactobacillus plantarum YS1 (L actinobacillus plantarum YS1) with a preservation number of CCTCC NO: M2016747.
2. Application of lactobacillus plantarum YS1 in preparation of food for preventing constipation.
The invention separates and identifies microorganisms in yak acid milk of Yushu Tibetan autonomous State, one of the Lactobacillus plantarum (L actinobacillus plantarum) is named as YS1, and the Lactobacillus plantarum is preserved in China center for type culture Collection (CCTCC for short, the address: Wuhan university No. eight 299 in Wuchang district, Wuhan City, Hubei province) in 2016, 12 months and 14 days, and the preservation number is CCTCC NO: M2016747.
The research results of an activated carbon-induced constipation model mouse show that lactobacillus plantarum YS1 has stronger gastric acid and bile salt resistance than lactobacillus bulgaricus, can inhibit the weight reduction of the mouse caused by constipation, reduce the weight, the number and the water content of excrement, simultaneously improve the propelling rate of activated carbon in small intestine, reduce the time for discharging first-particle black feces, and improve the serum level of motilin (MT L), gastrin (Gas), Endothelin (ET), acetylcholinesterase (AChE), Substance P (SP) and Vasoactive Intestinal Peptide (VIP) of a constipation mouse, and reduce the level of somatostatin (SS). the RT-PCR experimental results further show that lactobacillus plantarum YS1 can up-regulate the expression of c-Kit, SCF and GDNF mRNA in the intestinal tissue of the constipation mouse, down-regulate expression of TRPV1 and NOS mRNA.
The invention has the beneficial effects that: the invention provides lactobacillus plantarum YS1, which can effectively inhibit constipation, has an effect remarkably superior to that of commonly used lactobacillus bulgaricus, can be used for preparing food for preventing constipation, not only enlarges the application range of lactobacillus plantarum YS1 and improves the application value of the lactobacillus plantarum YS1, but also brings a new hope for preventing constipation.
Drawings
FIG. 1 is an agarose gel electrophoresis of genomic DNA of Lactobacillus plantarum YS1, wherein M is lambda DNA/Hind III Marker, and 1 is genomic DNA of Lactobacillus plantarum YS 1.
FIG. 2 shows the result of PCR amplification of 16S rDNA gene fragment of Lactobacillus plantarum YS1, where M is DNA Marker, C is negative control, and 1 is the product of PCR amplification of 16S rDNA gene fragment of Lactobacillus plantarum YS 1.
FIG. 3 shows the effect of Lactobacillus plantarum YS1 on charcoal-induced constipation in mice.
FIG. 4 shows the effect of Lactobacillus plantarum YS1 on charcoal-induced constipation of mice in the time to first-pellet-black stool excretion.
FIG. 5 is a graph showing the effect of Lactobacillus plantarum YS1 on charcoal-induced c-Kit and SCF mRNA expression in constipation mouse small intestine tissue.
FIG. 6 is a graph of the effect of Lactobacillus plantarum YS1 on charcoal-induced expression of TRPV1, GDNF, and NOS mRNAs in constipation mouse small intestinal tissue.
In FIGS. 3 to 6, L B represents Lactobacillus bulgaricus, and L P-YS1 represents Lactobacillus plantarum YS 1.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings.
Example 1 isolation and characterization of Lactobacillus plantarum YS1
Separation and purification of lactic acid bacteria
10 parts of naturally fermented yak yogurt samples collected from autonomous state herdsman families in Yushu Tibetan of Qinghai Yushu, each part of the sample is fully stirred uniformly by using a sterile glass rod, 50m L is sucked and put into a sterile centrifuge tube, the sterilized centrifuge tube is placed in a food sampling box and is taken back to a laboratory, and the sample is stored in a refrigerator at 4 ℃.
Taking 1m L sample of yak milk, and diluting with sterilized normal saline water by 10 times gradient to 10-7. Get 10-5、10-6、10-7Three dilutions of 1m L were filled with 15m L MRS medium (containing 5% CaCO by mass)3) The mixture is evenly mixed in a plate and is put in a constant temperature box with the temperature of 30 ℃ for culturing for 72 plus or minus 3 hours. Three replicates of each dilution were made. After the bacterial colony is formed, observing the morphological characteristics of the bacterial colony, selecting different bacterial colonies with calcium-dissolving rings, respectively inoculating the different bacterial colonies into a skim milk culture medium, culturing for 24-48 h at 30 ℃, streaking and inoculating the different bacterial colonies into an MRS agar culture medium after the bacterial strain grows well, and culturing for 48h at 30 ℃; the above steps were repeated at least three times until pure colonies were obtained.
Identification of Lactobacillus
Streaking pure strains on MRS solid culture medium, culturing at 30 deg.C for 48h, observing the apparent characteristics of colony size and shape with 10 times of magnifying glass, performing gram staining, and selecting strains with typical morphology for catalase test. Gram-positive and catalase test-negative coccal and bacillus strains were tentatively set as lactic acid bacteria.
The lactobacillus strains are subjected to growth temperature tests (10 ℃, 45 ℃, 60 ℃, 30min), pH value gradient tests, motility tests, catalase tests, oxidase tests, hydrogen sulfide tests, gelatin liquefaction tests, nitrate reduction tests, indole tests, glucose gas production tests, benzidine tests, litmus milk tests, arginine ammonia production tests, casein decomposition tests, various sugar alcohol fermentation tests and the like.
Results 20 lactobacillus strains are separated and purified from 10 yak milk samples, are gram-positive and catalase test negative, and are preliminarily identified as 7 lactobacillus strains through morphological observation and physiological and biochemical tests, wherein the number of the lactobacillus strains is L b1～Lb7, wherein the number is Lb4 was preliminarily identified as Lactobacillus plantarum (L Acobacterium plantarum) (Table 1).
TABLE 1 accession number Lb4 morphological characteristics and physiological and biochemical test results of Lactobacillus
Note: positive; -, negative; + w, weak positive.
Third, identification of Lactobacillus plantarum
Take the serial number Lb4, centrifuging to remove supernatant, collecting thallus, extracting genome DNA according to the method of a bacterial genome DNA extraction kit (Beijing Solebao scientific and technological company), performing agarose gel electrophoresis (the gel mass ratio concentration is 0.8%,voltage 80V, electrophoresis 80min), staining with GelRed nucleic acid dye, observing in gel imaging system (figure 1), diluting the extracted genome DNA by 200 times, determining concentration purity with ultraviolet spectrophotometer, performing PCR amplification on 16S rDNA gene fragment with primers 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' (SEQ ID No.1) and 1492R: 5'-GGCTACCTTGTTACGACTT-3' (SEQ ID No.2) (figure 2), sequencing the amplification product by bio-engineering (shanghai) limited and hokyo hexa-henhua Dacron technologies limited, and comparing the sequencing result (SEQ ID No.3) with the related sequence in GenbankbThe similarity between the 16S rDNA gene fragment sequence of lactobacillus 4 and L actinobacillus plantarum strain NBRC 15891 reaches 99%.
Hence, number Lb4, is identified as lactobacillus plantarum (L actinobacillus plantarum), is named as YS1, and is preserved in China center for type culture Collection (CCTCC for short, address: Wuhan university No. 299 in BaOnce district, Wuhan City, Hubei province, 12 months and 14 days in 2016), with the preservation number of CCTCC NO: M2016747.
Example 2 prevention of constipation in mice induced by activated charcoal with Lactobacillus plantarum YS1
In the early research on the constipation prevention effect of the lactic acid bacteria, the inventor proves that the defecation time of the first black stool, the levels of MT L, Gas, ET, SS, AChE, SP, VIP and the like can be used as indexes for evaluating the constipation prevention effect of the lactic acid bacteria, and the physiological effect of the lactic acid bacteria can be more exactly proved by detecting the expression of related mRNA in colon tissues through further molecular biological experiments.
The main materials, instruments and experimental animals used in this example were Lactobacillus plantarum YS1 (CCTCCNO: M2016747), Lactobacillus bulgaricus (China center for Industrial culture Collection of microorganisms), MT L, Gas, ET, SS, AChE, SP and VIP (Beijing Poir Biotech Co., Ltd.), Trizol reagent, OligodT18RNase, dNTP, M L V reverse transcriptionEnzymes (Invitrogen, usa); ROX reference Dye and SYBR Premix Ex Taq II (TAKARA, Japan); RT-PCR primers (Beijing Tiangen Biochemical technology Co., Ltd.). D550 digital cameras (canon, japan); a SevenEasy pH meter (mettler-toledo, switzerland); iMark microplate reader, T100 gradient PCR instrument (burle, usa). Seven-week-old female Kunming mice were purchased from Chongqing medical university laboratory animal center (license number: SYXK (Yu) 2012-0001), were raised in a temperature and humidity controlled environment (temperature 25 + -2 deg.C, relative humidity 50 + -5%), the light/dark cycle was adjusted every 12h, and standard feed and drinking water for the mice were provided.
First, detection of gastric acid resistance and bile salt tolerance of lactobacillus plantarum YS1
In order to exert probiotic action, lactic acid bacteria need to colonize and exert their physiological effects at the destination (usually the large intestine) under the strongly acidic conditions of the stomach and intestinal tract. Therefore, in order to research the potential probiotic effect of the lactic acid bacteria, an in vitro virtual model is established, and the detection of the gastric acid resistance and the bile salt tolerance of the lactic acid bacteria is an important detection means.
Dissolving NaCl and pepsin in distilled water, adjusting the pH value to 3.0 by using 1 mol/L of HCl to achieve the mass ratio of NaCl to 0.2%, adjusting the mass ratio of pepsin to 0.35%, filtering in vacuum to remove bacteria for later use as artificial gastric juice, activating lactobacillus plantarum YS1, culturing, centrifuging culture solution for 10min at the speed of 3000r/min for 5m L, collecting thalli, resuspending the thalli with 5m L of physiological saline, uniformly mixing with the artificial gastric juice according to the volume ratio of 1: 9, culturing at the temperature of 37 ℃ for 3h, measuring the viable count of lactobacillus plantarum YS1 at the time of 0h and 3h, and calculating the survival rate (%) of lactobacillus plantarum YS1 in the artificial gastric juice at the pH value of 3.0 by using a formula to be 3h (CFU/m L)/0 h (CFU/m L) × 100.
Inoculating Lactobacillus plantarum YS1 at an inoculation amount of 2% to MRS-THIO culture medium containing bilium bovis Seu Bubali salt at mass ratio of 0.0, 0.3%, 0.5%, and 1.0%, culturing at 37 deg.C for 24 hr, measuring absorbance at 600nm with blank culture medium of non-inoculated Lactobacillus plantarum YS1 as control, and calculating survival rate (%) of Lactobacillus plantarum YS1 in bile salt (medium OD) containing bile salt600Blank Medium OD600×100。
As shown in Table 2, the survival rate of Lactobacillus plantarum YS1 in artificial gastric juice at pH 3.0 and in bile salts at different concentrations was higher than that of Lactobacillus bulgaricus, and the survival rate of Lactobacillus plantarum YS1 in artificial gastric juice at pH 3.0 was about 2.5 times that of Lactobacillus bulgaricus, while the survival rate in bile salts at different concentrations was about 10 times that of Lactobacillus bulgaricus, indicating that Lactobacillus plantarum YS1 has stronger gastric acid resistance and bile salt tolerance than Lactobacillus bulgaricus.
TABLE 2 detection of gastric acid resistance and bile salt tolerance of Lactobacillus plantarum YS1
Secondly, prevention effect of lactobacillus plantarum YS1 on constipation induced by activated carbon in mice
100 Kunming mice with weight of about 25g are selected and averagely divided into 5 groups, namely a normal group, a constipation control group, a lactobacillus bulgaricus treatment group, a lactobacillus plantarum YS1 low-concentration treatment group and a high-concentration treatment group, after the mice in the normal group and the constipation control group are adapted to be raised for 1 week, the mice in the normal group and the constipation control group normally and freely take diet and drinking water for 2 weeks, and in the 2 weeks, the mice in the lactobacillus bulgaricus treatment group are respectively and daily gavage 2m L of 1.0 × 109CFU/m L Lactobacillus bulgaricus, Lactobacillus plantarum YS1 Low concentration treatment group and high concentration treatment group mice were each gavaged with 2m L1 × 10 per day8Lactobacillus plantarum YS1 and 1 × 10 of CFU/m L9After 1.2 weeks of lactobacillus plantarum YS of CFU/m L, 4 groups of mice were each gavaged with 2m L of activated carbon water (temperature 4 ℃, 10% activated carbon by mass was added to an aqueous solution containing 10% gum arabic to make a suspension) daily, while the groups were treated with lactic acid bacteria and the gavage was continued for 3 days, during the experiment, the body weight, dietary intake, water intake, feces weight and feces humidity of all the mice were measured daily at morning.
1. Effect of Lactobacillus plantarum YS1 on mouse body weight and feces
All mice were measured daily at morning intervals for the duration of the experiment for body weight, dietary intake, water intake, fecal weight and fecal moisture.
Body weight change is an important indicator of constipation in mice. As can be seen from fig. 3, the body weights of the mice of the groups in the first 2 weeks showed a normal growth state, and there was no significant difference between the groups; after constipation is induced by activated carbon, the weight of a normal group of mice which are not treated by the activated carbon continues to increase, and the weights of the rest groups of mice all decrease, wherein the weight of a constipation control group of mice decreases most, the weight of a lactobacillus plantarum YS1 high-concentration treatment group of mice is closest to that of the normal group and is obviously higher than that of a lactobacillus bulgaricus treatment group with the same concentration, which indicates that the lactobacillus plantarum YS1 can inhibit the weight decrease of the mice caused by constipation, and the effect is better than that of the lactobacillus bulgaricus.
The defecation state can show the constipation degree most obviously, the weight, the particle number and the water content of the excrement in the constipation state are important indexes of the excrement state, and the index decline shows the aggravation of the constipation degree. As shown in table 3, there were no significant differences in fecal weight, particle number, and water content between groups of mice on days 1-14; after constipation is induced by activated carbon, the weight, the particle number and the water content of the feces of the mice in a normal group are highest on 15-17 days, the control group with constipation is lowest, and the decrease of the weight, the particle number and the water content of the feces caused by constipation (P < 0.05) can be remarkably relieved by lactobacillus bulgaricus and lactobacillus plantarum YS1, but the weight, the particle number and the water content of the feces of the mice in a lactobacillus plantarum YS1 high-concentration treatment group are closest to those of the normal group and are remarkably higher than those of the lactobacillus bulgaricus treatment group with the same concentration, which shows that the lactobacillus plantarum YS1 can inhibit the decrease of the weight, the particle number and the water content of the feces of the mice caused by constipation, and the effect is better than that of.
TABLE 3 Effect of Lactobacillus plantarum YS1 on mouse faeces
Note: letter differences indicate significant differences between groups (P < 0.05).
2. Effect of Lactobacillus plantarum YS1 on activated carbon advancing rate and first-particle black feces excretion time
After the activated carbon water is infused into the stomach on the 17 th day, the mice are subjected to fasting for 24h, but the mice are still given free drinking water, after 24h, all the mice are infused with 0.2m L of activated carbon water per stomach, after 30min, half of the mice (10) in each group are subjected to neck-breaking and killed, the plasma of the mice is taken for standby, meanwhile, the intestines are taken to observe the propelling rate of the activated carbon in the intestines of the mice, the propelling rate (%) is the propelling length of the activated carbon/the total length of the intestines is × 100, and the rest 10 mice in each group continue to observe the time for discharging the first black feces.
Constipation causes a decrease in the frequency of intestinal peristalsis and an increase in the time during which feces stay in the intestine, causing the continuous reproduction of harmful bacteria using the feces as food, thereby threatening the health of the intestine and aggravating constipation. The length of the activated carbon propulsion and the rate of the activated carbon propulsion in the small intestine after the activated carbon induces the constipation of the mice can be used as indexes for evaluating the activity of the small intestine and the degree of constipation. As can be seen from Table 4, after constipation is induced by activated carbon, both Lactobacillus bulgaricus and Lactobacillus plantarum YS1 can improve the propelling length of activated carbon in the small intestine of a mouse, but the propelling rate of the activated carbon of the Lactobacillus plantarum YS1 high-concentration treatment group is the highest and is obviously higher than that of the Lactobacillus bulgaricus treatment group with the same concentration (P is less than 0.05) and the low-concentration treatment group of the Lactobacillus plantarum YS1, so that the Lactobacillus plantarum YS1 can improve the propelling rate of the activated carbon in the small intestine of the mouse, and the effect is better than that of the Lactobacillus bulgaricus.
TABLE 4 Effect of Lactobacillus plantarum YS1 on gastrointestinal propulsion in constipated mice
Note: letter differences indicate significant differences between groups (P < 0.05).
Constipation causes the slow movement of the intestinal tract, the retention time of the feces in the intestinal tract is increased, the discharge time of the first black feces is prolonged, and the shorter the discharge time of the first black feces means the more normal the movement of the intestinal tract. As can be seen from FIG. 4, the first black feces excretion time of the normal group was the shortest (88. + -.7 min), that of the constipation control group was the longest (231. + -.33 min), and that of the Lactobacillus plantarum YS1 high concentration treatment group was significantly shorter (132. + -.16 min) than that of the Lactobacillus plantarum YS1 low concentration treatment group (163. + -.22 min) and that of the Lactobacillus bulgaricus treatment group (175. + -.27 min) (P < 0.05), indicating that Lactobacillus plantarum YS1 could reduce the first black feces excretion time of the constipation mice, and had a good constipation relief effect, and the effect was superior to that of Lactobacillus bulgaricus.
3. Effect of Lactobacillus plantarum YS1 on mouse serum MT L, Gas, ET, SS, AChE, SP and VIP levels
The collected blood plasma of a mouse is taken, centrifuged for 15min at 4500rmp, serum is separated, MT L, Gas, ET, SS, AChE, SP and VIP levels of the blood serum of the mouse are measured according to the operation of a kit, MT L can stimulate the production of pepsin and promote intestinal tract movement, Gas can promote gastrointestinal secretion and gastrointestinal movement and promote pyloric relaxation to relieve constipation, ET plays an important role in vascular tension stability and basic cardiovascular system maintenance, SS can stimulate intestinal tract movement, AChE can condition muscle contraction and mucus secretion and can relax muscles to promote excrement discharge, SP is a substance which contributes to intestinal tract movement, and the normal content of VIP in the intestinal wall is also an important means for stabilizing the intestinal tract function.
As can be seen from Table 5, the serum of mice in the normal group had the highest MT L, Gas, ET, AChE, SP and VIP levels and the lowest SS level, while the mice in the constipation control group showed the opposite trend, and both Lactobacillus bulgaricus and Lactobacillus plantarum YS1 could make the serum levels of the above substances close to the normal group level, but the effect of Lactobacillus plantarum YS1 was more significant, indicating that Lactobacillus plantarum YS1 could make the serum MT L, Gas, ET, SS, AChE, SP and VIP levels of the constipation mice as normal as possible, thereby relieving constipation.
TABLE 5 Effect of Lactobacillus plantarum YS1 on mouse serum MT L, Gas, ET, SS, AChE, SP and VIP levels
Note: letter differences indicate significant differences between groups (P < 0.05).
4. Effect of Lactobacillus plantarum YS1 on mouse Small intestine tissue mRNA expression
Taking small intestine tissues of a mouse, crushing the small intestine tissues, extracting total RNA of colon tissues by using RNAzol, diluting the total RNA to 1 mu g/mu L, taking a total RNA extracting solution diluted by 2 mu L, sequentially adding oligodT18 of 1 mu L, RNase, dNTP, M L V enzyme and 5 × buffer of 10 mu L, synthesizing cDNA at 37 ℃ for 120min, 99 ℃ for 4min and 4 ℃ for 3min, then adopting the primers shown in the table 2, carrying out RT-PCR amplification on mRNA expression of c-Kit, SCF, TRPV1, GDNF and NOS, taking housekeeping gene GAPDH as internal reference, and finally carrying out agarose electrophoresis inspection on PCR amplification products by using ethidium bromide with the mass ratio of 1%.
TABLE 6RT-PCR primer sequences
ICC (interstitial cells of Cajal) in intestinal tracts of constipation patients is less, and c-Kit is a specific marker of ICC and is the key of ICC proliferation. The SCF concentration is important for the reproduction of ICC, which cannot grow in the absence of SCF. The literature reports that constipation mice have less ICC content in colon tissue and decreased c-Kit expression in colon tissue. The effect of lactobacillus plantarum YS1 on the expression of c-Kit and SCF mRNA in mouse small intestine tissue is shown in figure 5, the expression of c-Kit and SCF mRNA in the mouse small intestine of the constipation control group is reduced, lactobacillus plantarum YS1 can obviously increase the expression of c-Kit and SCF mRNA in the mouse small intestine, and the effect is more obvious at high concentration.
TRPV1 has been shown to be closely related to defecation and activation of TRPV1 can trigger the release of neurotransmitters, leading to small intestine dyskinesia. Increased expression of TRPV1 is a significant phenomenon of intestinal injury, and intestinal injury due to gastrointestinal disturbances causes higher expression of TRPV1 in constipation patients. GDNF can regulate the function of ganglion cells, thereby helping repair damaged intestines and preventing constipation. NOS plays an important role in regulating gastrointestinal motility. Increased NO can cause more severe colonic motility disorders, and control of NOS can reduce the content of NO, which is a feasible way to control constipation. The effect of lactobacillus plantarum YS1 on TRPV1, GDNF, and NOS mRNA expression in mouse intestinal tissue is shown in fig. 6, normal group mice had the highest GDNF mRNA expression and the lowest TRPV1, NOS mRNA expression, while constipation control group mice had the lowest GDNF mRNA expression, the highest TRPV1, and NOS mRNA expression, and both lactobacillus bulgaricus and lactobacillus plantarum YS1 could up-regulate GDNF mRNA expression and down-regulate TRPV1, NOS mRNA expression, relative to constipation control group, but lactobacillus plantarum YS1 was significantly better than lactobacillus bulgaricus.
Finally, it is noted that the above-mentioned embodiments illustrate rather than limit the invention, and that, while the invention has been described with reference to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.
<110> Chongqing second college of education; tanshanfang (Chinese character of 'Tanshan')
<120> Lactobacillus plantarum and application thereof in preparation of food for preventing constipation
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<210>11
<211>17
<212>DNA
<213> Artificial sequence
<220>
<223> downstream primer for RT-PCR amplification of GDNF mRNA expression
<400>11
agcccaaacc caagtca 17
<210>12
<211>21
<212>DNA
<213> Artificial sequence
<220>
<223> upstream primer for amplifying NOS mRNA expression by RT-PCR
<400>12
ccacatctgg caggatgaga a 21
<210>13
<211>21
<212>DNA
<213> Artificial sequence
<220>
<223> downstream primer for amplifying NOS mRNA expression by RT-PCR
<400>13
aggcacagaa ctgagggtac a 21
<210>14
<211>20
<212>DNA
<213> Artificial sequence
<220>
<223> upstream primer for amplification of GAPDH mRNA expression by RT-PCR
<400>14
cggagtcaac ggatttggtc 20
<210>15
<211>20
<212>DNA
<213> Artificial sequence
<220>
<223> downstream primer for amplification of GAPDH mRNA expression by RT-PCR
<400>15
agccttctcc atggtcgtga 20
Claims (2)
1. Lactobacillus plantarum YS1(Lactobacillus plantarumYS1), the preservation number is CCTCC NO: m2016747.
2. Use of lactobacillus plantarum YS1 according to claim 1 for the preparation of a food product for the prevention of constipation.
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| CN107099491A (en) * | 2017-07-05 | 2017-08-29 | 重庆第二师范学院 | Lactobacillus plantarum YS3 and its application in the food of Constipation is prepared |
| CN107267416A (en) * | 2017-07-05 | 2017-10-20 | 重庆第二师范学院 | Lactobacillus plantarum YS2 and its application in the food of Constipation is prepared |
| CN107723256B (en) * | 2017-09-01 | 2020-11-06 | 杭州娃哈哈科技有限公司 | Lactobacillus plantarum new strain and application thereof |
| CN109700033B (en) * | 2019-03-04 | 2021-06-04 | 山东环亿生物科技有限公司 | Probiotic composition for regulating intestinal tract and preventing constipation and preparation method thereof |
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| EP2937424B1 (en) * | 2014-04-23 | 2017-03-29 | National Yang-Ming University | Lactic acid bacterium, composition containing the same and their use |
| CN104498383B (en) * | 2014-05-28 | 2017-12-26 | 西南大学 | A kind of lactobacillus fermenti Lactobacillus fermentum strain suo and application thereof of adjustable intestinal movement, Constipation |
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Non-Patent Citations (1)
| Title |
|---|
| Improvement of constipation and liver function by plant-derived lactic acid bacteria: A double-blind, randomized trial;Fumiko Higashikawa et al.;《Nutrition》;20101231;第26卷;第367-374页 * |
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Effective date of registration: 20230606 Address after: 201799 No. 10666, Songze Avenue, Qingpu District, Shanghai Patentee after: Biogrowing. Co.,Ltd. Address before: No.9, Xuefu Avenue, Nan'an District, Chongqing, 400067 Patentee before: CHONGQING University OF EDUCATION Patentee before: Tan Fang |