![]() Two factors are important for the function of sgRNA promoter activity and the specificity of the sgRNA. 2013), then directs the Cas nuclease to cleave the target sequence. SgRNA cassette: sgRNA is a programmable 20-nucleotide (nt) sequence that recognizes the target DNA sequence and an invariant scaffold sequence (Ran et al. The expression of Hpt is usually driven by a strong constitutive promoter, such as maize Ubiquitin 1 ( ZmUbi1), rice ACTIN 1, or Cauliflower mosaic virus ( CaMV) 35S, for the ubiquitous expression of the antibiotic-tolerance gene (Mikami et al. Plant phosphomannose isomerase can also be used as a selectable marker for rice transformation (Hu et al. Among these genes, Hpt is the most widely used selection marker which confers tolerance to the herbicide hygromycin because several crops have a natural tolerance to kanamycin. Selection cassette: In monocots, several genes have served as useful selection markers for the efficient selection of transgenic plants, such as neomycin- phosphotransferase ( NPTII), bar, mutated acetolactate synthase ( ALS), plant phosphomannose isomerase, and hygromycin phosphotransferase ( Hpt) (Miki and McHugh 2004 Hu et al. Plasmids used for the stable genome editing of plants require a selection cassette, known as a sgRNA cassette, and a clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) cassette in the T-DNA region, while the selection cassette is not necessary for transient transformations (Fig. For transient transformations, particle bombardment and polyethylene glycol (PEG)-mediated methods are used to deliver plasmids or ribonucleoproteins (RNPs) into the plant cells (Woo et al. For stable transformations, the Agrobacterium-mediated transformation method is typically used to deliver transfer DNA (T-DNA) into the plant cell, where it is then inserted into the plant genome (Mikami et al. Genome-edited plants can be generated either by stable or transient transformation. To perform CRISPR/Cas9-mediated genome editing, the Cas9 endonuclease is guided by a single guide RNA (sgRNA) to recognize the complementary sequence and create double-strand breaks (DSBs), thereby generating a short deletion or insertion. The CRISPR/Cas9 system has been successfully used for genome editing in a variety of monocots, including rice ( Oryza sativa), wheat ( Triticum sp.), barley ( Hordeum vulgare), and maize ( Zea mays) (Feng et al. We have aimed to encompass a full spectrum of information for genome editing in monocot crops. Four topics are covered: the design and construction of plasmids for genome editing in monocots alternatives to SpCas9 protoplasts and CRISPR and screening for marker-free CRISPR/Cas9-induced mutants. In this review, we introduce new techniques for genome editing and identifying marker-free genome-edited mutants in monocot crops. RNPs can be used to efficiently generate transgene-free genome-edited crops that can reduce transgene issues related to the generation of genetically modified organisms. Moreover, the delivery of Cas9-sgRNA ribonucleoproteins (RNPs) is one strategy that can be used to prevent transgene issues with the expression of sgRNA and Cas proteins. Before the stable transformation of a plant, the evaluation of vectors and target sites is therefore very important. Furthermore, to reduce the sequence limitation of the protospacer adjacent motif (PAM) and for other purposes, many Cas protein variants and base editors can be used in plants. Currently, various vectors and online tools are available to aid sgRNA design. Analyses of successful targeted mutations have revealed that the expression levels, expression timing, and variants of both sgRNA and Cas9 need to be sophisticatedly regulated therefore, the promoters of these genes and the target site positions are the key factors for genome-editing efficiency. To generate genome-edited crops, single guide (sg)RNA and Cas9 DNA are delivered into plant cells and expressed, and the predicted position is targeted. The breakthrough CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9-mediated genome-editing technology has led to great progress in monocot research however, several factors need to be considered for the efficient implementation of this technology.
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![]() Kaleidoscope stars Giancarlo Esposito ( The Mandalorian season 3, Breaking Bad), Jai Courteney ( The Suicide Squad), Paz Vega (Rambo: Last Blood), Rufus Sewell (Old), Tati Gabrielle (Uncharted), and Peter Mark Kendall (The Americans) among others. That way, Kaleidoscope will keep you guessing right up until its last moments and force you to rethink what you know about its puzzle pieces with each passing episode. The idea is that every subscriber will get a unique viewing experience, with the order in which they watch Kaleidoscope's episodes affecting how they view each crew member, when the plot is going, and which questions they'll have (and answers they'll get) after each installment. The only episode that every viewer will end on is White, which marks the epic conclusion to the story. Once that episode is complete, you'll get another episode to watch, such as Violet (set 24 years before the heist) or Pink (six months post-heist). Meanwhile, your best friend might kick off their adventure with the episode titled Red, which starts on the day of the heist itself. For example, Netflix might start you off with the entry called Yellow, which is set six weeks before the heist. A ray incident on one mirror at angle, θ after reflection falls.So, how will Kaleidoscope work? When you begin watching the series, you'll start with a specific episode (randomly assigned to you by Netflix), each one of which is named after a particular color – hence the title Kaleidoscope. Two plane mirrors are inclined at 70 ∘.The critical angle for glass-water boundary is C.The phenomenon involved in the reflection of radio-waves by ionosphere is similar to?.Two plane mirrors are inclined to each other ray of light incident on one face at 30 ∘ after reflection…?.A ray of light from a denser medium strikes a rarer medium at an angle of incidence I.A cord attached to a vibrating tuning fork is divided into six segments under tension…?.One of its refracting surfaces is silvered. An object is placed at a distance 20cm from the pole of a convex mirror of focal length…?.Yellow light is used in a single slit diffraction experiment with slit width of 0.6 mm.A wave has velocity u in medium P and velocity 2u in medium Q.Two plane mirrors A and B are aligned parallel to each other, as shown in the figure.White light is incident on the interface of glass and air as shown in the figure.This will be through the multiple reflections of the single slice appearing side by side. Now, if you place the slice of pizza within two mirrors that are kept adjacent to each other at an angle, you will be able to see almost the entire pizza. In order to understand what happens in a kaleidoscope, imagine a pizza. Each slice of the pizza depicts the object put within the triangular area at the edge of the tube. Place the second triangle upside down so that there is enough space for plastic to move between the two transparencies.Īnd, once you are done with the making of kaleidoscope, you can design it beautifully with glitters, colours and so on. Attach another triangular transparent paper that is larger than the first one. Step 7: Place the colored plastic pieces at the end of the kaleidoscope that has transparent paper. Take some small paper cuttings that can be placed on the transparent paper. Step 6: Select the kind of plastic colour you would want to have inside the kaleidoscope. Step 5: Draw another triangle, but this time make it 2 cm larger than the earlier one. Step 4: Now, fix the paper into a separate place. Cut thin openings at the corners for easier folds. Step 3: After that, place the transparent paper at the edge of the Kaleidoscope. Step 2: Sketch the small triangle that is present at the edge of the kaleidoscope to the transparency paper (keep 1 cm extra around the triangle for allowing folding of the paper). Make sure that it is solid and taped to the exterior of the triangle. Step 1: Get three mirror pieces and roll them in the form of a triangle. Modern Item Weight, 27 pounds Product Dimensions, 31.5 x 21 x 17 inches ASIN. ![]() These instruments are an ideal choice for progressing young students in need of an instrument that can meet their developing needs through high school, college, and often, beyond. Yamaha YEP-321S 4-valve Intermediate Euphonium - Silver-plated Style. Step-Up Instruments (also called Intermediate Instruments) are designed with the professional features an advancing player needs, but on an instrument that can be purchased at a manageable price. Yamaha YEP-321 Euphonium 4 +8 Bb Euphonium 4 Périnet valves Bell: 280 mm Bore: 14.5 - 15. Why Step-Up? As a musicians' skills become more advanced, their instruments need to do the same. We take the time to inspect & address every item so that the instrument you receive is not just brand new - it's better than brand new. All band instruments (from even the finest brands) can be subject to small inconsistencies & manufacturing residues that can affect how the instrument plays. The 321 features a strong warm tone, and is easy to play, with accurate intonation. Why Heid? All band instruments at Heid Music are professionally inspected & approved for quality by the esteemed Heid Music Repair Team. Yamaha YEP-321 Series 4-Valve Euphonium - Lacquer. 36-Month Financing Open quick view dialog for Allora AEP-450 Vienna 4-Valve Euphonium. Or 76 /month with 36 month financing Limited Time. Laser-fused bell is plasma welded to create a seamless brass medium for your sound vibrations to flow through 4.5 of 5 stars (25 Reviews) Or 76 /month with 36 month financing Limited Time. Perfectly rounded pressure formed tubing to encourage smoother air flow Vysvtlivky k nastavení tisku: - Zvolte VELIKOST - zde si zvolíte pesnou velikost jedlého papíru jaká vám a potebám vaeho dortu vyhovuje. Honed valves precisely fit to eliminate air leakage Includes 48M mouthpiece and hardshell case. Combine those attributes with its 4 quick-action nickel-plated top valves and you've got the perfect combination to create some serious euphonium tone. Its small shank receiver allows for focused air flow, easy tone production, and excellent response. The silver-plated Yamaha YEP-321S Euphonium is a favorite of advancing students and band directors, as well as brass professionals looking to double on the euphonium. I'm ready to start my new student rental!. As the fresh sheet of paper replaced the printed paper, the now freshly inked rollers ran over the type again. Meanwhile, a sheet of paper slid against a hinged platen (see image), which then rapidly pressed onto the type and swung back again as the sheet was removed and the next sheet inserted. With the advent of industrial mechanisation, inking was carried out by rollers that passed over the face of the type, then moved out of the way onto an ink plate to pick up a fresh film of ink for the next sheet. At right are several small platen jobbing presses, at left, a cylinder press. Ink rollers made of composition made inking faster and paved the way for further automation.ġ917 press room, using a line shaft power system. Later metal presses used a knuckle and lever arrangement instead of the screw, but the principle was the same. Printers' workshops, previously unknown in Europe before the mid-15th century, were found in every important metropolis by 1500. This form of presswork gradually replaced the hand-copied manuscripts of scribes and illuminators as the most prevalent form of printing. At 1,282 pages, it took him and his staff of 20 almost 3 years to complete. It was Gutenberg's "screw press" or hand press that was used to print 180 copies of the Bible. Gutenberg also invented a wooden printing press, based on the extant wine press, where the type surface was inked with leather-covered ink balls and paper laid carefully on top by hand, then slid under a padded surface and pressure applied from above by a large threaded screw. Johannes Gutenberg is credited with the development in the western hemisphere, in about 1440, of modern movable type printing from individually cast, reusable letters set together in a "forme" (frame or chase). ![]() Movable type was first invented in China using ceramic type in 1040 AD during the Northern Song dynasty by the inventor Bi Sheng (990–1051). Printer operating a Gutenberg-style screw press More recently, letterpress printing has seen a revival in an artisanal form. Letterpress printing remained the primary means of printing and distributing information until the 20th century, when offset printing was developed, which largely supplanted its role in printing books and newspapers. Letterpress printing was the normal form of printing text from its invention by Johannes Gutenberg in the mid-15th century to the 19th century and remained in wide use for books and other uses until the second half of the 20th century. thick between the bed and the paper can be printed using letterpress. In theory, anything that is "type high" and so forms a layer exactly 0.918 in. ![]() With certain letterpress units, it is also possible to join movable type with slugs cast using hot metal typesetting. In practice, letterpress also includes wood engravings, photo-etched zinc plates ("cuts") linoleum blocks, which can be used alongside metal type wood type in a single operation stereotypes and electrotypes of type and blocks. A worker composes and locks movable type into the "bed" or "chase" of a press, inks it, and presses paper against it to transfer the ink from the type, which creates an impression on the paper. Letterpress printing is a technique of relief printing for producing many copies by repeated direct impression of an inked, raised surface against individual sheets of paper or a continuous roll of paper. The general form of letterpress printing with a platen press shows the relationship between the forme (the type), the pressure, the ink, and the paper. ![]() We are committed to ensuring that your information is secure. We may use the information to customise the website according to your interests. We may contact you by email, phone, fax or mail. From time to time, we may also use your information to contact you for market research purposes.We may periodically send promotional emails about new products, special offers or other information which we think you may find interesting using the email address which you have provided.We may use the information to improve our products and services.We require this information to understand your needs and provide you with a better service, and in particular for the following reasons: What we do with the information we gather other information relevant to customer surveys and/or offers.demographic information such as postcode, preferences and interests.contact information including email address.We may collect the following information: You should check this page from time to time to ensure that you are happy with any changes. We may change this policy from time to time by updating this page. 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The client will report satisfactory pain control at a level of less than 3 to 4 on a scale of 0 to 10.The client will use pharmacological and nonpharmacological pain relief strategies.Spasms in the lower back and bladder area. ![]() Inflammation and infection of the urinary tract (e.g., urethra, bladder, and other urinary tract structures). ![]() Burning or pain when urinating is one of the common symptoms of UTI (Urology Care Foundation, 2022). The irritation causes pain in the lower abdomen or pelvic area and even in the lower back. The lining of the bladder becomes inflamed and irritated. Complicated UTIs are defined as UTIs that are associated with metabolic disorders, that is secondary to anatomic or functional abnormalities that impair urinary tract drainage, or that involve unusual pathogens, which increase the risk for therapeutic failure (Brusch & Stuart, 2023).īecause of the referred pain pathways, even simple lower UTI may be accompanied by flank pain and costovertebral angle tenderness. An uncomplicated UTI usually only involves the bladder. Bacteria may ascend to the ureters to multiply and cause infection of the kidneys ( pyelonephritis). An infection in the urethra is called urethritis. UTIs are usually classified as infections involving the upper or lower urinary tract. These traits facilitate colonization and infection of the urinary tract (Brusch & Stuart, 2023). Uropathogenic bacteria, derived from a subset of fecal flora, have traits that enable adherence, growth, and resistance to host defenses. But, in some cases, bacteria overcome the natural defenses of the body, therefore causing infection. Usually, bacteria that enter the urinary tract system are removed by the body before they can cause symptoms. Among the pathogens responsible for the remainder are Staphylococcus saprophyticus, Proteus mirabilis, Klebsiella pneumoniae, or Enterococcus faecalis. E.coli causes the majority of uncomplicated cystitis cases. coli), normally found in the digestive system. ![]() Most UTIs are caused by the bacterium Escherichia coli ( E. These infections account for a significant number of emergency department (ED) visits, and 20% of women develop at least one UTI (Brusch & Stuart, 2023). UTI is defined as significant bacteriuria in the setting of symptoms of cystitis or pyelonephritis. Urinary tract infections (UTIs) are caused by pathogenic microorganisms in the urinary tract (kidney, bladder, urethra). Discover the evidence-based nursing interventions for urinary tract infection in this guide. Get to know the nursing diagnosis for urinary tract infection (UTI) nursing care plans. ![]() He’s so often depicted as a greedy hoarder who raises taxes for his own Scrooge McDuck-level treasury, but history tells us these taxes went to the “Saladin tithe”, a ten-percent tax increase to fund King Richard’s Crusade. What happens when you acknowledge heroism in your enemies? Why can’t the Sheriff of Nottingham be a noble leader, trying desperately to defend his good citizens from the wanton criminals surrounding his city? What actions might he take- at the sacrifice of his own reputation -to protect Nottingham from domestic terrorism? Because it’s impossible not to empathize with someone once you sit down with them and discover they’re (gasp!) an actual human. In Star Wars, we’ve seen exactly one Imperial Stormtrooper take his helmet off, and he instantly became one of the heroes of the movie. In NOTTINGHAM I strove to make the members of the Sheriff’s Guard as varied and personable as the Merry Men. Instead of an endless army of faceless guards, let’s take the helmets off and learn their names. But rather than accept this villainy at face value, let’s undo the dehumanization. Whereas a villain has a tiny circle of Us, which is why they’re cruel to anyone who doesn’t serve their personal needs.Īnything that shrinks your radius of Us moves you closer to being the bad guy.ĭoes this apply to the Sheriff of Nottingham? For centuries we’ve been told he’s greedy and evil, and that his hordes of minions are heartless killers. Heroes have a very large circle of Us, which is why they’re willing to make personal sacrifices to benefit their larger community. Look back at our defining characteristics of heroes and villains. If you’ve ever instinctively dismissed someone because of their political party, generational identity, or geography, then you (or more likely, somebody who is manipulating you) have indeed just become the bad guy. We see this every day, especially in partisan bickering that paints the opposing side with broad brushstrokes. Is this starting to sound like politics? It should. ![]() So are we really the bad guy, or have we just been unfairly dehumanized? In order to bolster your Us against your Them, you have to make Them a threat. Which means that, from their perspective, we’re the bad guy. We like to think our enemies are selfish because it makes them easier to hate, when in reality we’re simply not inside their circle of Us. The Sheriff of Nottingham, on the other hand, was a real person. Seems pretty simple-so string up your bow, it’s time to do hero stuff!īut while impossibly cruel villains work great in sci-fi/fantasy genres (it’s how we get Negans, Voldemorts, and Saurons), they don’t usually ring as realistic. If you’re Robin Hood, you probably drew a circle around “the people.” And you probably think the Sheriff drew a circle just barely big enough to fit himself and his comically unwieldy bags of money. Especially if there’s a sword within reach. And if something outside that circle threatens something inside the circle, you know what to do. Voila! With one simple scribble, you’ve created your own personally-tailored line between good guys and bad guys. Here’s a simple exercise: Take every person and thing you care about and draw a circle around them. Identity-stifling costumes is an easy trick for the movies, but in real life we have to dehumanize them emotionally. “Non-human”? “Zero value on their lives?” That’s right, the key to labeling people as evil is to dehumanize them, so we don’t have to fuss with those messy emotions we’d feel if we had to care about them. We don’t flinch when heroes have to dispatch them by the dozen. This generally works fine for baddies such as aliens, robots, zombies, Nazis, and orcs they’re so inarguably evil and non-human that we literally place zero value on their lives. Whether they’re medieval guards, post-apocalyptic scavengers, or Imperial Stormtroopers, there’s always an army of infinitely-killable minions battling our hero. ![]() It’s easy to pinpoint “bad guys” in movies: they’re usually dressed nearly identically, and you often can’t even see their faces. After all, does anyone really consider themselves “the bad guy”? WHAT EXACTLY IS A BAD GUY? ![]() But how do you believably write from an evil mindset? Many stories have been told “from the villain’s perspective,” but these overwhelmingly rely upon a series of sympathetic events that drive a character to become the villain (think the Star Wars prequel trilogy) rather than truly seeing things from their point of view. In my historical epic Nottingham I’ve chosen to break those traditional black-and-white roles and follow perspectives on both sides. ![]() Go to our main page for more Porn Comics. She did not know what was happening and tried to hide the pleasure and her prayers in the middle of the class so that nobody noticed anything. Blackfire (real name Komandr) is the firstborn eldest daughter and princess of the long-dead King and Queen of Tamaran and the older sister of Starfire and. She started sucking and fucking her while Raven was in class. He decides to open a portal in the book where Raven’s sweet pussy could be seen. 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Cover the hair with the included hair cap and process no longer than 20 minutes.Gently work and massage product into the hair to ensure it is evenly saturated.Wearing gloves, apply liberally and quickly to clean and dry hair, focusing on color to be removed.Shake well for 30 seconds and, pointed away from your face, cut tip on applicator cap.Add contents from bottle #1 into applicator bottle #2.*Not formulated for direct-application dyes as well as bold shades such as pinks, blues, greens, purples, etc. Best used to create a clean slate allowing you to easily re-color hair or transition to a new shade. IDEAL FOR: Hair that has been colored with a regular, oxidative (mixed with developer) dye. This bleach-free and ammonia-free process means you can safely re-color your hair the same day. See Reconstructor page for full details.Ĭolor Oops Extra Strength Hair Color Remover is an easy and effective way to reverse undesirable hair color by shrinking the dye molecules, allowing you to simply wash them away. ![]() MUST ADD RECONSTRUCTOR TO CART TO RECEIVE FREE GIFT WHILE SUPPLIES LAST. FREE RECONSTRUCTOR CONDITIONER WITH PURCHASE OF EXTRA STRENGTH OR EXTRA CONDITIONING HAIR COLOR REMOVER (TWO MAX). ![]() In contrast, an agrarian society's daily routines for work and personal conduct are more likely governed by the length of daylight hours and by solar time, which change seasonally because of the Earth's axial tilt. ![]() The time of day that individuals begin and end work or school, and the coordination of mass transit, for example, usually remain constant year-round. Industrialized societies usually follow a clock-based schedule for daily activities that do not change throughout the course of the year. A minority of the world's population uses DST Asia, Africa, and Latin America and the Caribbean generally do not.Īn ancient water clock that lets hour lengths vary with season. The United States observes it, except for the states of Hawaii and Arizona (within the latter, however, the Navajo Nation does observe it, conforming to federal practice). Conversely, it is not observed at some places at high latitudes, because there are wide variations in sunrise and sunset times and a one-hour shift would relatively not make much difference. Some countries observe it only in some regions: for example, parts of Australia observe it, while other parts do not. DST is generally not observed near the Equator, where sunrise and sunset times do not vary enough to justify it. Many countries have used DST at various times since then, particularly since the 1970s energy crisis. Starting on 30 April 1916, the German Empire and Austria-Hungary each organized the first nationwide implementation in their jurisdictions. In 1908, Port Arthur in Ontario, Canada, now merged into Thunder Bay, started using DST. After some serious consideration, it was not implemented. In 1907, British resident William Willett presented the idea as a way to save energy. ![]() In 1895, New Zealand entomologist and astronomer George Hudson proposed the idea of changing clocks by two hours every spring to the Wellington Philosophical Society. In a satirical letter to the editor of The Journal of Paris, Franklin suggested that waking up earlier in the summer would economize on candle usage and calculated considerable savings. The idea of aligning waking hours to daylight hours to conserve candles was first proposed in 1784 by U.S. As a result, there is one 23-hour day in early spring and one 25-hour day in the middle of autumn. The typical implementation of DST is to set clocks forward by one hour in either the late winter or spring (" spring forward"), and to set clocks back by one hour in the fall (" fall back") to return to standard time. Daylight saving time ( DST), also referred to as daylight savings time, daylight time (United States, Canada, and Australia), or summer time (United Kingdom, European Union, and others), is the practice of advancing clocks (typically by one hour) during warmer months so that darkness falls at a later clock time. |