CN109897850A - DNA method is extracted in biological sample release on forensic lignocellulosic carrier - Google Patents
DNA method is extracted in biological sample release on forensic lignocellulosic carrier Download PDFInfo
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- CN109897850A CN109897850A CN201910136032.7A CN201910136032A CN109897850A CN 109897850 A CN109897850 A CN 109897850A CN 201910136032 A CN201910136032 A CN 201910136032A CN 109897850 A CN109897850 A CN 109897850A
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Abstract
The invention discloses biological sample releases on a kind of forensic lignocellulosic carrier to extract DNA method, vehicle treated enzymatic hydrolysis solution is made using cellulase, pectase, zytase compatibility, it degrades under the conditions of pH7 ~ 11 to the cellulose carrier containing biological sample DNA, to make biological sample efficiently separate with carrier fibre, biological sample high-quality inside carrier fibre is largely released from the carrier being degraded, DNA is improved and extracts quality and effect；The present invention establishes a kind of utilization vehicle treated enzyme, and degradation has the lignocellulosic carrier of biological evidence, to realize that biological sample is released effectively to extract the new method of DNA.
Description
Technical field
The invention discloses biological sample releases on a kind of forensic lignocellulosic carrier to extract DNA method, establishes
A kind of to utilize vehicle treated enzyme, degradation has the lignocellulosic carrier of biological evidence, to realize that biological sample is released effectively
To extract the new method of DNA, belong to forensic identification technology field.
Technical background
In forensic medicine in appraisal of material evidence field, the carrier of biological sample is saved based on lignocellulosic, such as blood collection card, saliva
Capture card, cotton swab, cotton, denim, books, paper, plank (stick), stalk etc..Due to save during dehydration and
The dry preservation state in cool place so that biological sample is combined closely with lignocellulosic, can protect in this way biological sample not by
Degradation, and long-term preservation.But as the holding time extends, covalent cross-linking can occur between biological sample and lignocellulosic, especially
Its biological sample crosslinking for being in fibrous outer surfaces is more serious, so that can not usually be obtained using existing DNA extraction method
Good DNA inspection result.
For DNA extraction angle, forensic DNA extraction method mainly has organic solvent method, polystyrene two at present
Vinyl benzene resin method, silica gel method, paramagnetic particle method, cetyl trimethylammonium bromide method, pellosil absorption method, ethanol precipitation purifying
The combination of method, isopropanol precipitating method of purification and these methods, these methods are all made of three key steps, it may be assumed that 1, take it is suitable
Amount is enclosed with the carrier of biological sample, carries out integral hydrolysis or enzymatic hydrolysis to biological sample using different physico chemical factors or protease；
2, the DNA in biological sample is discharged into solution；3, the DNA that recovery purifying or direct use are discharged into solution is for subsequent inspection
It surveys.As can be seen that these methods have an obvious deficiency, that is, it is intended only to start with from biological sample itself and carrier surface, passes through
Solution acts on from outside to inside, and digestion cracks biological sample protein or smudge cells to obtain DNA, and for being embedded in close load
Then undertreatment, biological sample can not fill the biological sample and cellulose in internal portion-biological sample crosslinking inner face biological sample
Point release so that the holding time is long, albuminous degeneration degree is serious, and with the serious biological sample of carrier crosslinking, it is existing
Biological sample processing and DNA extraction method are all undesirable, even can not obtain effective dna sometimes, thus waste valuable life
Object sample and live sample, affect the acquirement of cracking of cases and key evidence.
It is well known that DNA save it is best, influenced the smallest part by extraneous factor and be all present in carrier inside or fiber
Element-biological sample inner surface, but still release this part biological sample sufficiently without method at present, one of reason is just
It is the absence of the method that biological sample is released from carrier and carries out DNA extraction.
Summary of the invention
The invention discloses biological sample releases on a kind of forensic lignocellulosic carrier to extract DNA method (below
Abbreviation vehicle treated enzyme extraction method), vehicle treated enzymatic hydrolysis solution is made using cellulase, pectase, zytase compatibility,
It degrades under the conditions of pH7 ~ 11 to the cellulose carrier containing biological sample DNA, so that biological sample be made to have with carrier fibre
Effect separation largely releases biological sample high-quality inside carrier fibre from the carrier being degraded, and improves DNA and extracts matter
Amount and effect.
DNA method is extracted in biological sample release on a kind of forensic lignocellulosic carrier provided by the invention, including
Following steps:
1) the cellulose family carrier with biological sample is sheared into 1mm2 ~ 10cm2；
2) 0.01 ~ 300mg/ml of cellulase final concentration, 0.01 ~ 300mg/ml of pectase final concentration, zytase final concentration are taken
0.01 ~ 300mg/ml is added in the buffer of pH7 ~ 11, mixes well, vehicle treated enzyme solutions are made；
3) biological sample sheared is put into containing in 0.005 ~ 10ml enzymatic hydrolysis solution, soaks biological sample by enzyme solution completely
Not yet, 40 DEG C ~ 55 DEG C heated at constant temperature of water-bath react 0.1 ~ 10 hour, sufficiently enzymatic hydrolysis cellulose carrier；
4) reaction solution is sucked out after directly carrying out DNA extraction inspection or 5000 ~ 12000rpm of reaction solution being centrifuged 3min, in removal
Clearly, sediment carries out DNA and extracts inspection.
It is described with lignocellulose carrier include: blood collection card, saliva capture card, cotton swab, cotton, denim,
Books, paper, plank (stick), stalk etc..
Biological sample includes: blood (spot), saliva (spot), sweat (spot), sperm (spot), urine (spot), tear (spot), group
Knit liquid (spot) etc..
In addition, the method for the present invention can also be used for other with lignocellulose carrier and lignocellulose biological sample
Extract detection.
Since it is difficult to extract DNA from the biological sample for being embedded in lignocellulosic carrier inside, STR-PCR inspection is affected
It tests as a result, therefore, the present invention is to breach traditional concept, creatively utilizes cellulase, pectase, zytase compatibility system
At vehicle treated enzyme solutions, start with from lignocellulose degradation class carrier, under the conditions of pH7 ~ 11, make inside cellulose carrier and
The high-quality biological sample of cellulose-biological sample inner surface is sufficiently discharged, and is obtained better DNA and is extracted inspection result.This
It is more significant that invention extracts DNA effect to the release containing micro-biological sample in the biological sample, carrier of preservation for a long time.
The positive effect of the present invention is: for forensic mostly using lignocellulose material as carrier the characteristics of, it is sharp
With vehicle treated enzyme lignocellulose degradation carrier, with the skin gone, what can the hair adhere to, and it is new to establish biological sample release extraction DNA
Method enables carrier inside to save best biological sample and sufficiently discharges, and improves forensic dna and extracts quality and effect.This
In invention, vehicle treated enzyme uses the buffer system of pH7 ~ 11, reduces the damage of DNA in extraction process.The result shows that the present invention
The biological sample DNA on lignocellulose carrier can be effectively extracted, accurate to obtain STR-PCR parting information, effect is better than
Other existing DNA extraction methods.
Detailed description of the invention
Fig. 1 is 1 control group electron microscope of embodiment；
Fig. 2 is that embodiment 1 utilizes vehicle treated enzymatic treatment carrier cotton electron microscope；
Fig. 3 is the STR parting map for the blood collection card that embodiment 2 is saved 11 years using the method for the present invention processing；
Fig. 4 is the STR parting map for the blood collection card that embodiment 2 is not saved using present invention processing 11 years；
Fig. 5 is the STR parting map for the saliva capture card that embodiment 3 is saved 9 years using the method for the present invention processing；
Fig. 6 is the STR parting map for the saliva capture card that embodiment 3 is not saved using present invention processing 9 years；
Fig. 7 is the STR parting map that embodiment 4 saves 17 years bloodstain cotton swabs using the method for the present invention processing；
Fig. 8 is the STR parting map that embodiment 4 does not save 17 years bloodstain cotton swabs using present invention processing；
Fig. 9 is the STR parting map that embodiment 5 saves 13 years bloodstain cottons using the method for the present invention processing；
Figure 10 is the STR parting map that embodiment 5 does not save 13 years bloodstain cottons using present invention processing；
Figure 11 is the STR parting map that embodiment 6 has bloodstain newspaper for 3 years using the method for the present invention processing preservation；
Figure 12 is the STR parting map that embodiment 6 does not save 3 years newspapers with bloodstain using present invention processing.
Specific embodiment.
To facilitate the understanding of the present invention, especially exemplified by following embodiment.Its act on be understood to be to explaination of the invention and
It is non-to any type of limitation of the invention.
Embodiment 1
In order to which checking carrier treatment enzyme can do the experiment of this embodiment with degraded cellulose carrier.Prepare cellulose carrier processing
Cellulase (1mg/ml), pectase (2mg/ml), zytase (5mg/ml) are added into the phosphate buffer of pH8 for enzyme solution,
Final volume is 1ml.0.4cm2 blood collection card is put into cellulose carrier processing enzyme solution, is totally submerged it, as experiment
Group.In addition, 0.4cm2 blood collection card is put into 1ml phosphate buffer, as a control group.By experimental group and 45 DEG C of control group
Water-bath 2 hours, 8000 revs/min of centrifugations took supernatant respectively, and DNS method surveys reduced sugar.The results show that in experimental group, reduced sugar
Concentration is 0.1mg/ml, and reduced sugar is not detected in control group.From the experimental results, vehicle treated enzyme can effectively degrade
Cellulose carrier.
Blood collection card after the degradation of vehicle treated enzyme is done into scanning electron microscopic observation.As shown, Fig. 1 is control group Electronic Speculum
Photo, Fig. 2 are experimental group electromicroscopic photograph.In Fig. 1, fiber is cylindric, smooth surface；In Fig. 2, fiber surface is uneven
It is whole, there is apparent recess, shows that vehicle treated enzyme has apparent degradation to fiber.
Embodiment 2
11 years blood collection card clip 0.4cm2 for having bloodstain will be saved.Prepare cellulose carrier processing enzyme solution (pH8): fiber
Plain enzyme final concentration 1mg/ml, pectase final concentration 2mg/ml, zytase final concentration 5mg/ml.The 0.4cm2 blood sheared is adopted
Truck is respectively put into the EP pipe containing 0.4ml vehicle treated enzyme and 0.4ml physiological saline (control), is totally submerged it, water
45 DEG C of heating are bathed to react 2 hours.After 5000 rpm are centrifuged 3min, supernatant is removed, 0.5%SDS, 2ul Proteinase K is added in sediment
(10mg/ml), TNE buffer, 37 DEG C keep the temperature 0.5 ~ 2 hour, supernatant be added isometric phenol, etc. ratio mixed phenol and chlorine
After imitative, chloroform respectively extracts once, supernatant is taken to add NaCl the or NaAc solution of 1/10 volume and the cold dehydrated alcohol of 2.5 times of volumes
It mixing, 10000rpm centrifugation 2min removes supernatant, and the mixing of 70% ethyl alcohol is added in precipitating, and 10000rpm centrifugation 2min removes supernatant,
TE buffer is added after drying or pure water is spare.
It is expanded using 21 system of Powerplex, the DNA extracted is added according to the proportion that kit requires
PCR instrument is put into after adding to be expanded.Get out 96 orifice plates of sequencing and the sample expanded.The sample number being sequenced as required
10ul formamide containging interior traget and suitable corresponding amplified production is added in equal number of holes in amount, another independent
It is loaded ladder 1ul, as a result as shown in Figure 3 and Figure 4.From in Fig. 3, Fig. 4 as can be seen that in routine inspection (control) sample
PCR amplification unbalanced phenomena is obvious, and 4 locus such as PentaD, PentaE, CSF1PO, TPOX do not obtain parting, D5S818,
There is allelic loss in D13S317 locus, and vehicle treated enzyme extraction method obtains the complete of whole 21 str locus seats
Parting, amplification unbalanced phenomena are obviously improved, illustrate to obtain more good DNA using vehicle treated enzyme extraction method, DNA is mentioned
It takes effect more preferable, is conducive to STR-PCR and examines.
Embodiment 3
9 years saliva capture card clip 0.25cm2 with salivary stain will be saved.Prepare cellulose carrier processing enzyme solution (pH9):
Cellulase final concentration 2mg/ml, pectase final concentration 4.5mg/ml, zytase final concentration 10mg/ml.The saliva that will be sheared
Capture card is respectively put into the EP pipe containing 0.35ml vehicle treated enzyme and 0.35ml physiological saline (control), soaks it completely
Not yet, 50 DEG C of heating of water-bath are reacted 1.5 hours.After 5000rpm is centrifuged 3min, supernatant is removed, 150ul polystyrene diethyl is added in sediment
Alkenyl benzene resin (5%) and 2ul protease k(10mg/ml), it is put into 56 DEG C of digestion 30min of metal bath.After digestion, it is put into 98 DEG C of gold
Belong to and be denaturalized 10 ~ 15min in bath, 12000rpm is centrifuged 2min, places 4 DEG C of refrigerators and save backup.
It is expanded using 21 system of Powerplex, the DNA extracted is added according to the proportion that kit requires
PCR instrument is put into after adding to be expanded.Get out 96 orifice plates of sequencing and the sample expanded.The sample number being sequenced as required
10ul formamide containging interior traget and suitable corresponding amplified production is added in equal number of holes in amount, another independent
It is loaded ladder 1ul, as a result as shown in Figure 5 and Figure 6.As can be known from Fig. 5 and Fig. 6, vehicle treated enzyme extraction method obtains
All 21 correct typing datas of str locus seat, no allelic loss, uneven amplification, the peak Pull up, stutter with,
Phenomena such as peak split, occurs, and in Routine control experiment, 4 locus such as D13S317, PentaE, PentaD, TPOX do not obtain
STR parting is obtained, 3 locus such as D2S1338, CSF1PO, D5S818 allelic loss occur, illustrate and control experiment phase
Than extracting saliva capture card sample using vehicle treated enzyme extraction method and obtaining the higher DNA of quality, DNA extraction effect is better than
Routine control experiment.
Embodiment 4
The cotton swab clip with bloodstain for saving 17 years is appropriate.Prepare cellulose carrier processing enzyme solution (pH9): cellulase is whole
Concentration 6mg/ml, pectase final concentration 7mg/ml, zytase final concentration 10mg/ml.The bloodstain cotton swab sheared is put into respectively
In EP pipe containing 0.3ml vehicle treated enzyme and 0.3ml physiological saline (control), 50 DEG C of heating of water-bath are reacted 1 hour.
Supernatant is removed after 5000rpm centrifugation 3min, 150 ul polystyrene divinylbenzene resins (5%) and 2ul protease is added in sediment
K(10mg/ml), it is put into 56 DEG C of digestion 30min of metal bath.After digestion, it is put into 10 ~ 15min of denaturation in 98 DEG C of metal baths,
12000rpm is centrifuged 2min, places 4 DEG C of refrigerators and saves backup.
It is expanded using 21 system of Powerplex, the DNA extracted is added according to the proportion that kit requires
PCR instrument is put into after adding to be expanded.Get out 96 orifice plates of sequencing and the sample expanded.The sample number being sequenced as required
10ul formamide containging interior traget and suitable corresponding amplified production is added in equal number of holes in amount, another independent
It is loaded ladder 1ul, as a result as shown in Figure 7 and Figure 8.As can be seen that being mentioned using vehicle treated enzyme extraction method from Fig. 7, Fig. 8
Cotton swab bloodstain DNA is taken to obtain all 21 accurate partings of str locus seat, it is harmonious more preferable, and control experiment PentaE, TPOX
Locus is not detected, and amplification unbalanced phenomena is obvious, illustrates the cotton swab bloodstain DNA mass of vehicle treated enzyme extraction method extraction more
It is good, more it is able to satisfy STR-PCR amplification needs.
Embodiment 5
13 years cotton clip 0.25cm2 with bloodstain will be saved, prepares cellulose carrier processing enzyme solution (pH8): cellulose
Enzyme final concentration 6mg/ml, pectase final concentration 7mg/ml, zytase final concentration 10mg/ml.The bloodstain cotton that will be sheared respectively
It is put into EP pipe and 0.3ml physiological saline (control) containing 0.3ml vehicle treated enzyme, 55 DEG C of heating of water-bath are reacted 2 hours.
After 5000rpm is centrifuged 3min, supernatant is removed.Appropriate lysate, 95 ~ 100 DEG C of 30min of metal bath are added in sediment.5000rpm centrifugation
Precipitating is removed after 3min, supernatant is added magnetic bead liquid, is sufficiently mixed, room temperature 5min.Vortex vibrates on postposition magnet stand, removes liquid
Body.On addition lysate, cleaning solution oscillation washing postposition magnet stand, liquid is removed, is air-dried 5 ~ 10min at room temperature, is added appropriate pure
Water, 65 DEG C of 5min.Postposition magnet stand is mixed, DNA solution is sucked out spare.
It is expanded using 21 system of Powerplex, the DNA extracted is added according to the proportion that kit requires
PCR instrument is put into after adding to be expanded.Get out 96 orifice plates of sequencing and the sample expanded.The sample number being sequenced as required
10ul formamide containging interior traget and suitable corresponding amplified production is added in equal number of holes in amount, another independent
It is loaded ladder 1ul, as a result as shown in Figure 9 and Figure 10.It can be seen from the figure that in control experiment (Figure 10), PentaE,
3 locus such as PentaD, TPOX are not detected, and allelic loss occurs in D5S818 locus, and vehicle treated enzyme extraction method
Complete 21 str locus seat parting is obtained after the DNA cloning of acquisition, parting is accurate, expands harmonious more preferable, no allele
It loses, the DNA purity for illustrating that vehicle treated enzyme extraction method extraction cotton bloodstain obtains is higher, and integrality is more preferable.
Embodiment 6
3 years newspaper clip 0.3cm2 with bloodstain will be saved.Prepare cellulose carrier processing enzyme solution (pH9): cellulase
Final concentration 6mg/ml, pectase final concentration 7mg/ml, zytase final concentration 10mg/ml.The bloodstain newspaper sheared is put respectively
Enter in the EP pipe containing 1.5ml vehicle treated enzyme and 1.5ml physiological saline (control), be totally submerged sample, 40 DEG C of water-bath add
Thermal response 1 hour.After 5000rpm is centrifuged 3min, supernatant is removed, suitable 150ul lysate and 2ul Proteinase K is added in sediment
It after (10mg/ml), is uniformly mixed, 56 DEG C keep the temperature 1 hour, 85 DEG C of 10min of metal bath.Supernatant mistake is taken after 12000rpm centrifugation 3min
Ethanol on pellosil post purifying, adds pure water to elute, eluent is spare.
It is expanded using 21 system of Powerplex, the sample extracted is carried out according to the proportion that kit requires
PCR instrument is put into after addition to be expanded.Get out 96 orifice plates of sequencing and the sample expanded.The sample number being sequenced as required
10ul formamide containging interior traget and suitable corresponding amplified production is added in equal number of holes in amount, another independent
It is loaded ladder 1ul, as a result as is illustrated by figs. 11 and 12.It can be seen from figure 11 that vehicle treated enzyme extraction method obtains whole
21 str locus seat partings, as a result accurately, no allelic loss, uneven amplification, the peak Pull up, stutter with,
Phenomena such as peak split, occurs, although and control experiment has also detected full gene seat, overall peaks it is obvious it is relatively low (<
400rfu), the DNA quantity for illustrating that vehicle treated enzyme extraction method obtains is significantly more than control experiment.
Embodiment 7
20 preservations 10 years or more the blood collection cards for having blood cake are taken, respectively clip 0.8cm2.Formulation vehicle handles enzyme solution,
In include: cellulase final concentration 2mg/ml, pectase final concentration 4.5mg/ml, zytase final concentration 10mg/ml, with pH8
Buffer mix well, obtain vehicle treated enzyme solution.The bloodstain capture card sheared is respectively put into containing 1.5ml vehicle treated
In the EP pipe of enzyme and 1.5ml physiological saline (control), 55 DEG C of heating of water-bath are reacted 1 hour.After 5000rpm is centrifuged 3min, go
Clearly, appropriate 5% polystyrene divinylbenzene resin and 2ul protease k(10mg/ml is added in sediment), it is put into 56 DEG C of metal bath and disappears
Change 30min.After digestion, it is put into 10 ~ 15min of denaturation in 98 DEG C of metal baths, 2min is centrifuged not less than 10000rpm, places 4 DEG C of ice
Case saves backup.
Table 1, vehicle treated enzyme process and polystyrene divinylbenzene resin method extract DNA result comparison sheet
It is expanded, i.e., added the sample extracted according to the proportion that kit requires using 21 system of Powerplex
PCR instrument is put into after adding to be expanded.Get out 96 orifice plates of sequencing, and the sample expanded, the sample number being sequenced as required
10ul formamide containging interior traget and suitable corresponding amplified production is added in equal number of holes in amount, another independent
It is loaded ladder 1ul.
The result shows that (such as table 1), using vehicle treated Enzymatic Extraction DNA, expanding effect and the correct parting gene of acquisition
Seat quantity is substantially better than polystyrene divinylbenzene resin method, especially obtains the sample size of 19 ~ 21 locus, the present invention
The case of method is 3 times for not using case of the present invention, and the DNA mass for illustrating that the method for the present invention obtains is substantially better than polystyrene
Divinylbenzene resin method.
Claims (5)
1. a kind of by the vehicle treated enzyme solutions that biological sample discharges on lignocellulosic carrier in forensic, feature
It is mainly to be prepared by following raw material:
Cellulase: 0.01 ~ 300mg/ml；
Pectase: 0.01 ~ 300mg/ml；
Zytase: 0.01 ~ 300mg/ml.
2. DNA method is extracted in biological sample release on a kind of forensic lignocellulosic carrier as described in claim 1,
It is characterized in that:
It takes cellulase, pectase, zytase to be added in the buffer of pH7 ~ 11 by above-mentioned concentration, mixes well, load is made
Body handles enzyme solutions.
3. DNA method is extracted in biological sample release on a kind of forensic lignocellulosic carrier, comprising the following steps:
1) the lignocellulose carrier with biological sample is sheared into 1mm2 ~ 10cm2；
2) vehicle treated enzyme solutions are prepared by method for claim 2；
3) biological sample sheared is put into the enzymatic hydrolysis solution of 0.005 ~ 10ml step 2 preparation, makes biological sample quilt completely
Enzyme solution submergence, 40 DEG C ~ 55 DEG C heated at constant temperature of water-bath react 0.1 ~ 10 hour, sufficiently enzymatic hydrolysis cellulose carrier；
4) reaction solution is sucked out after directly carrying out DNA extraction inspection or 5000 ~ 12000rpm of reaction solution being centrifuged 3min, in removal
Clearly, sediment carries out DNA and extracts inspection.
4. as claimed in claim 3 a kind of by biological sample carries out release extraction on lignocellulosic carrier in forensic
The method of DNA, it is characterised in that:
It is described with lignocellulose carrier include: blood collection card, saliva capture card, cotton swab, cotton, denim, books,
Paper, plank (stick), stalk etc.；
Biological sample includes: blood (spot), saliva (spot), sweat (spot), sperm (spot), urine (spot), tear (spot), tissue fluid
(spot) etc..
5. as claimed in claim 3 a kind of by biological sample carries out release extraction on lignocellulosic carrier in forensic
The method of DNA can be used for other and be detected with the extraction of lignocellulose carrier and lignocellulose biological sample.
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| CN101631502A (en) * | 2006-12-29 | 2010-01-20 | 恰根有限公司 | Trigger the method and the material that discharge biological sample |
| WO2012000888A1 (en) * | 2010-06-29 | 2012-01-05 | Dsm Ip Assets B.V. | Polypeptide having acetyl xylan esterase activity and uses thereof |
| CN102439145A (en) * | 2009-04-24 | 2012-05-02 | 帝斯曼知识产权资产管理有限公司 | Carbohydrate degrading polypeptide and uses thereof |
| CN102834511A (en) * | 2010-02-11 | 2012-12-19 | 帝斯曼知识产权资产管理有限公司 | Polypeptide having cellobiohydrolase activity and uses thereof |
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2019
- 2019-02-25 CN CN201910136032.7A patent/CN109897850A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101631502A (en) * | 2006-12-29 | 2010-01-20 | 恰根有限公司 | Trigger the method and the material that discharge biological sample |
| CN102439145A (en) * | 2009-04-24 | 2012-05-02 | 帝斯曼知识产权资产管理有限公司 | Carbohydrate degrading polypeptide and uses thereof |
| CN102834511A (en) * | 2010-02-11 | 2012-12-19 | 帝斯曼知识产权资产管理有限公司 | Polypeptide having cellobiohydrolase activity and uses thereof |
| WO2012000888A1 (en) * | 2010-06-29 | 2012-01-05 | Dsm Ip Assets B.V. | Polypeptide having acetyl xylan esterase activity and uses thereof |
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