Insect pathogenic fungus interacts with the gut microbiota to. The following protocol is provided for the purification of dna from yeast and filamentous fungi. Qiagen gentra puregene blood kit 30 ml, for 30 ml blood. The v3 and v4 variable regions of the 16s rrna gene were amplified and sequenced on the illumina miseq platform. Gentra puregene blood kit 30 ml 158445 from qiagen sample. Vortex using light pulses except in step 7 where thorough vortexing is key. Total dna was prepared using the qiagen gentra puregene yeast bact kit with overnight cultures of a. Gentra puregene blood kit from qiagen selectscience. Broth medium oxoid, uk and inc u bated overnight at. Complete genome sequence of brevundimonas diminuta atccb. Shotgun library preparation was performed using a truseq nano dna kit with a target insert size of 350bp illumina, san diego, ca. Kg supplemented with yeast extract, dna was extracted and purified using the gentra puregene yeast bact. Gentra puregene yeast bact kit gramnegative protocol.
A complete solution for sampletostorage purification. Samples using the gentra puregene blood kit this protocol is for purification of genomic dna from 10 ml compromised whole blood using the gentra puregene blood kit. We next focused manual annotations on gene families with. Qiagen gentra puregene blood kit 120 ml, for 120 ml blood. Kit for purification of archivequality dna from yeast and grampositive or gramnegative. For purification of archivequality dna from 200 ml bacterial cultures or 400 ml yeast cultures. Materials and methods sample setup oragene kits were used to collect 2 ml saliva samples from 16 individual donors following the instructions for collection provided with the kits. Equivalent amounts of yeast or bacteria were processed using the zr fungalbacterial dna kit or the kit from competitor m. Product lines include puregene r and versagene rna and dna purification kits as well as the autopure lsr and autotechtm instruments. Genomic dna for longread sequencing was extracted from overnightgrown cells using the gentra puregene yeast bact kit protocol qiagen, hilden, germany. Biomass was ground by alternating beadbeating for 4. Dna purification from yeast using the gentra puregene yeastbact. Spherical body protein 2 truncated copy 11 as a specific babesia bovis attenuation marker article snippet. Complete or highquality draft genome sequences of six.
Total genomic dna was extracted from 1 ml of liquid culture with the gentra puregene yeast bact. The gentra puregene tissue kit enables purification of highmolecularweight 100200 kb dna suitable for archiving. Kit qiagen using a modified protocol, in which all vortexing steps were replaced by mixing with gentle pipetting to maintain the integrity of replication intermediates. Genomic dna from 10 diverse strains pna 971r, pna 0601, pna 2003, pna 151, pna 997, pans 0402, pans 0102, pans 9923, pans 9936, and pans 993 was obtained from overnight cultures started from single colonies using the gentra puregene yeast bact kit qiagen inc. Gentra puregene blood kit 30 ml from qiagen selectscience. The manual improvement of the genomes was finalized with bandage ver. Gentra puregene qiagen dna isolation carl baker, university of washington, 40114 1. Jul 17, 2007 i have routinely used the puregene kit for isolation of genomic dna from mouse tails in order to screen transgenic and knockout mouse colonies with pcr and southern blots. Ten micrograms of purified dna was processed with the pacific biosciences pacbio 10kbp library kit in the nc state university genomic sciences laboratory and then analyzed by singlemolecule realtime smrt rs ii sequencing. Gentra puregene blood kit 120 ml 158467 from qiagen. Cell suspension solution, lytic enzyme solution, rnase a solution, and reagents.
The following steps were performed according to the protocol. Archivequality dna for longterm storage reproducible dna purification a complete solution for sampletostorage purification convenient, scalable purification procedure gentra puregene blood. The reads were assembled by the newbler program, which generated 121 contigs. Gentra puregene blood kit ml, 158389 from qiagen labsave. Sequencing of am202, am368, am854, am936, am1041, and am1096 a gentra puregene kit qiagen, valencia, ca was used to extract dna from the blood of cattle infected with the following strains and isolates of a. Header goes here in title case and should be no longer. The genomic dna was extracted with gentra puregene yeast bact. For nanopore sequencing, genomic dna was extracted using a gentra puregene yeast bact. Draft genome sequences of three isolates of golubevia sp. Thisisolatedgdnawasusedfordeep sequencing, quantitative realtime pcr qpcr studies, and human oral microbe identi. Dc3000 genomic dna ngl prepared using the gentra puregene yeast bact.
Equipment and reagents to be supplied by user when working with chemicals, always wear a suitable lab coat, disposable gloves, and protective goggles. Genomic library preparation and sequencing were outsourced to macrogen seoul, south korea. Puregene mousetail kit from gentra now qiagen biocompare. Cell suspension solution, lytic enzyme solution, rnase a solution, and reagents archivequality dna for longterm storage. Draft genome sequences of two glycoalkaloiddegrading. For purification of archivequality dna from yeast and grampositive or gramnegative bacteria. Rnase a solution, puregene proteinase k, and reagents. Jun 06, 2017 at each of five time points 0, 12, 36, 60, and 84 h after fungal infection, mosquitoes were dissected to collect midguts. Kit 49 dna purification from grampositive bacteria using the gentra puregene yeast bact. Insect pathogenic fungus interacts with the gut microbiota.
Kit is intended for molecular biology applications. Kit for purification of archivequality dna from yeast and grampositive or gramnegative bacteria. Compatibility of the puregene dna purification kit with the. Dna was isolated with the gentra puregene yeast bact. Masterpure yeast dna purification kit manual lucigen. Ten micrograms of purified dna was processed with the pacific biosciences pacbio 10kbp library kit in the nc state university genomic sciences. Gentra puregene tissue kit for purification of archivequality dna from tissues. To prevent degradation of dna, avoid vigorous vortexing and heating. The scalable purification procedure gently removes contaminants and inhibitors and allows samples to be purified for use as longterm references. For mouse tail screening, qiagen who recently purchased gentra now provides kits for isolation of 100 mg and 4 g of mouse tails. The remainder of gdna isolation followedthemanufacturersprotocol.
Dna extraction patient whole blood collected in purpletop tube was received at the indiana university molecular genetics diagnostic laboratory, indianapolis, in, and dna extracted using the qiagens gentra puregene blood kit qiagen, germantown, md following the. We extracted genomic dna from wasps with a puregene dna extraction kit gentra, minneapolis, usa. Catalytically inactive cas9 impairs dna replication fork. This product is not intended for the diagnosis, prevention, or treatment of a disease. Qiagen gentra puregene blood kit for purification of archivequality dna from whole blood, packed cells, buffy coat, and bone marrow features of the qiagen puregene blood kit include. Simple, manual shearing method using 26g needles for fragmentation of gdna 30 kb. Shop now at fisher scientific for all of your scientific needs. It can isolate dna from grampositive and gramnegative bacteria, fungi, algae, and. Closed genome sequences of clinical neisseria gonorrhoeae. Kit 200 ml 158667 gentra puregene tissue kit 4 g 158689 gentra puregene tissue kit 33 g 158745 gentra puregene cell kit 2 x 108.
The sequencing library was prepared using a truseq dna pcrfree library preparation kit and sequenced with a hiseq 2000 platform illumina, usa, with pairedend read lengths of 101bp. Simple, manual shearing method using 26g needles for fragmentation of gdna 30 kbdna solution is passed through the syringe needle multiple times e. Rapid nanopore sequencing of plasmids and resistance gene. Intragenic cftr duplication and 5t12tg variant in a patient with nonclassic cystic fibrosis article snippet. Kit enables purification of highmolecularweight 100200 kb dna suitable for archiving. Gentra puregene blood kit for purification of archivequality dna from whole blood, packed cells, buffy coat, and bone marrow. Genome sequence of aeromicrobium erythreum nrrl b3381, an. The bacterial dna was purified using gentra puregene yeast bact. Pacbio americas user group meeting sample prep workshop. Prevalence of streptococci and increased polymicrobial.
Library preparation for wholegenome sequencing was performed using the nextera dna library prep kit illumina, inc. The dna extracts obtained with three different dna extraction protocols gentra puregene yeast bact kit grampositive protocol. Required fields by submitting this form, you submit your information to qiagen, who will use it to communicate with you regarding this topic. Genomic dna from the prototype strain pna 971r was obtained from overnight cultures started from single colonies using the gentra puregene yeast bact kit qiagen inc, quantified using biospectrometer eppendorf, and visually assessed for quality on a 0. Determination of concentration, yield, purity, and length of dna 61.
For purification of archivequality dna from whole blood, packed cells, buffy coat, and bone marrow. Qiagen may also use your personal information for internal business and marketing purposes, including but not limited to webinar and raffle registration, product, trial, educational and events information. Genomic landscape and genetic manipulation of the black. Kansas 6de, dawn australia, colville c51 and c52, kansas em, nayarit, mexico n3518, n4506, and n3571. Dna gdna extraction using the gentra puregene yeast bact kit qiagen. Kit enables purification of highmolecular weight 100200 kb dna suitable for archiving. For subsequent purification of the dna, the nucleospin genomic dna cleanup kit machereynagel was used. Browse a full range of gentra systems products from leading suppliers. Gentra systems is a leading provider of nucleic acid purification technology, instrumentation and innovation to the global life sciences industries. Equipment and reagents to be supplied by user when working with chemicals, always wear a suitable lab coat, disposable gloves, and protective.
Do not heat samples except when doing the rnase a step. After revival on nutrient yeast extract glycerol agar plates 4 for 2 days. Quickdna fungalbacterial miniprep kit zymo research. Genome sequence of in vitro probiotic strain isolated from a. Genome sequence of the marine bacterium vibrio campbellii. Qiagen genomictip 20100500g kit qiagen gentra puregene yeast bact. Samples were then incubated at 50c overnight in an air incubator. Rbc lysis solution, rnase a solution, and reagents. Dna extract quantity and purity was varied betwee n the different extraction kits and a260a280 and. Frontiers pantoea ananatis genetic diversity analysis. M of each dntp, 200 nm of each primer, 1 ng template dna and 0.
Genomic dna was purified from liquid cultures grown at 20c in lysogenic broth lb using the gentra puregene yeast bact. For purification of archivequality dna from human whole blood, bone marrow, buffy coat, buccal cells, body fluids, cultured cells, tissue, mouse tail, yeast, bacteria. The dna concentration was quantified and quality controlled using a nanodrop 2000 instrument thermo scientific, rockford, il and qubit fluorometer life technologies, grand island, ny. Determination of concentration, yield, purity, and length of dna. Equal volumes of eluted dna were then analyzed in a 0. Qiagen gentra puregene tissue kit 4 g, for 4 g tissue. For purification of archivequality dna from human whole blood, bone marrow, buffy coat, buccal cells, body fluids, cultured cells, tissue, mouse tail, yeast, bacteria pdf 367kb. Gentra systems puregene chemistries from the entire 4 ml sample contained in the oragene kit. Microbial dna was prepared from midguts using the gentra puregene yeast bact kit b qiagen, following.
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