![]() In this paper, we have adapted the clustered regularly interspersed palindromic repeats (CRISPR) system from Streptococcus pyogenes to repress genes in trans in the cyanobacterium Synechococcus sp. Trans-acting regulators provide novel opportunities to study essential genes and regulate metabolic pathways. Our combinatorial method using a Cas9 nickase genome editing tool to integrate the gene repression module demonstrates easy-to-use and high-efficiency advantages, paving the way for stably manipulating genes, even essential ones, for functional characterization and microbial engineering. These results indicate the effectiveness of tandem promoter-driven RNA repression modules in promoting gene repression in chromosomal integrants. With the design of a tandem repetitive promoter-driven asRNA module and the use of a new Cas9 nickase genome editing tool, a chromosomal integrant (LM3P) was generated in a single step and successfully enhanced RNA repression, with a 27% decrease in acetate titer on cellulose in antibioticfree medium. However, further testing on chromosomal integrants that were created by double-crossover recombination exhibited only very weak repression because DNA integration dramatically lessened gene dosage. The strains with pta gene repression generally diverted more carbon into ethanol. To control both acetate and more » lactate simultaneously, we transformed the repression plasmid into lactate production-deficient mutant and found the plasmid delivery reduced acetate titer by more than 33%, concomitant with negligible lactate formation. Our plasmid-based expression of antisense RNA (asRNA) molecules targeting the phosphotransacetylase (pta) gene successfully reduced the enzymatic activity by 35% in cellobiose-grown cells, metabolically decreased the acetate titer by 15 and 52% in wildtype transformants on cellulose and xylan, respectively. Here we developed a Cas9 nickase-assisted chromosome-based RNA repression to stably manipulate essential genes in C. Clostridium cellulolyticum is a promising candidate for consolidated bioprocessing however, its genetic manipulation to reduce the formation of less-valuable acetate is technically challenging due to the essentiality of acetate-producing genes. ELSAs enable simultaneous and stable regulation of many genes for metabolic engineering and synthetic biology = ,Įssential gene functions remain largely underexplored in bacteria. Using ELSAs, we created three highly selective phenotypes in Escherichia coli, including redirecting metabolism to increase succinic acid production by 150-fold, knocking down amino acid biosynthesis to create a multi-auxotrophic strain, and repressing stress responses to reduce persister cell formation by 21-fold. ![]() ![]() We design ELSAs by combining non-repetitive genetic parts according to algorithmic rules quantifying DNA synthesis complexity, sgRNA expression, sgRNA targeting, and genetic stability. We apply biophysical modeling, biochemical characterization, and machine learning to develop toolboxes of non-repetitive genetic parts, including 28 sgRNA handles that bind Cas9. We stably co-expressed 22 sgRNAs within non-repetitive extra-long sgRNA arrays (ELSAs) to simultaneously repress up to 13 genes by up to 3500-fold. When using CRISPR interference, co-expressing many single-guide RNAs (sgRNAs) triggers genetic instability and phenotype loss, due to the presence of repetitive DNA sequences. Engineering cellular phenotypes often requires the regulation of many genes.
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Now that you've mapped out your product plan, your team will want to know: who's doing what when? This is where a FigJam product development roadmap template is especially handy, with built-in tools to tag individual team members, set team priorities, and assign key tasks.
![]() With Alpena County, MI's population of 28,929, there is a high likelihood you will find weed from a close friend, or a person in your legal cannabis social network. Additionally, cannabis has sixty other cannabinoids to research and provide symptom relief. Any plant that produces a pleasant psychoactive drug called tetrahydrocannabinol (THC) from its over four hundred fifty compounds is a true gift. Marijuana, also known as cannabis among other names and slangs, like "weed", has never really been that hard to find for medical or recreational use. Cannabis Facts and Information Alpena County, MI Consumers and Usage Information It is also safe if the Alpena County dispensary has a valid Michigan retail cannabis license approved for delivery. In fact, it's like any other home delivery service experience. Don't worry, ordering marijuana delivery from a licensed business is very similar to ordering pizza. Finally, they take payment in cash or card and the Alpena County dispensaries delivery order is complete.Ĭannabis brings with it a host of emotions and opinions due to many years of negative propaganda and reefer madness. The delivery person checks for valid identification. When the delivery time arrives, a delivery person brings cannabis to your home. First, you find a dispensary and marijuana menu on the Internet. It is a similar process in areas where Alpena County dispensaries are legal. Meaning, a person with cannabis products comes to a home and weed is provided for a cash payment. Not surprisingly, the weed delivery experience is lot like the black market most have experienced. Finally, ensure that weed delivery is legal in the intended delivery city of Alpena County. Next, find out if the legal Alpena County dispensaries are approved for delivery. The most important thing a consumer can do first is ensure that cannabis for purchase is legal in Michigan dispensaries. When you purchase cannabis for delivery from a licensed marijuana business, it is safe. Any business activity or purchase from a black market dispensary is a state and federal crime. If you see advertising for Alpena County dispensaries that are not listed here, assume the business is black market operating illegally. Businesses listed here are legal and operating with a valid Michigan state cannabis license. To know if you can purchase weed legally in Alpena County dispensaries or any city within Michigan, check our listings. Since marijuana is new in terms of legalization, the different county and city scenarios are many. Other cities may only allow marijuana production or lab-testing businesses. Some cities will allow marijuana delivery but not permit dispensaries to operate a storefront in a community. Some cities ban cannabis sales even though the state has legalized it. Alpena County, Michigan is not unique and these types of issues are present. Unfortunately, all this does is fuel a local Alpena County dispensaries black market and hinder the legal market efforts. Some counties ban delivery of cannabis products thinking they will keep marijuana out. Unlike public residents voting on issues at the state level, a city or county's leaders may vote to ban marijuana dispensaries and other cannabis related businesses. US states, counties, and cities are all different organizations or corporations and leaders can make their own cannabis rules. Share Tweet Is it Legal to Buy Weed at Alpena County Dispensaries?Īs with most cannabis legality questions, understanding whether you can buy weed at Alpena County dispensaries is complex. ![]() Tmesis, the use of a lexical word rather than an affix, is sometimes considered a type of infixation. Examples: itíc 'did s/he plant it?' and iti tóoc 'did they sow it?'. In Seri, some verbs form the plural stem with infixation of ⟨tóo⟩ after the first vowel of the root compare the singular stem ic 'plant (verb)' with the plural stem itóoc. The word 'gigi' means "tooth", while 'g erigi' means "serration"'.The word 'cerlang' means "luminous", while 'c emerlang' means "brilliant"'.The word 'gembung' (variant of 'kembung') means "bloated", while 'g elembung' means "bubble"'.All infixes are no longer productive and cannot be used to derive new words. In Malay and Indonesian, there are three infixes ( sisipan), ⟨el⟩, ⟨em⟩, and ⟨er⟩. These elements are no longer productive, and occur crystallized in words inherited from Old Khmer. They include the nominalizing infix ⟨b⟩, which derives l bɨən 'speed' from lɨən 'fast' and l bɑɑng ' trial' from lɔɔng 'to test, to haunt', or the agentive ⟨m⟩ deriving c mam 'watchman' from cam 'to watch'. Khmer, an Austroasiatic language, has seven different infixes. Tagalog has borrowed the English word graduate as a verb to say "I graduated" a speaker uses the derived form gr umaduate. The most common infix is ⟨in⟩ used to make an intentional verb, as in ' giniba', meaning ‘ruined’ (from ‘ giba’, an adjective meaning ‘worn-out’) ' binato’, meaning ‘stoned’ (from ‘ bato’, ‘stone’) and ' ginamit’, meaning ‘used’. For example, in Tagalog, a grammatical form similar to the active voice is formed by adding the infix ⟨um⟩ near the beginning of a verb. Infixes are common in Austronesian and Austroasiatic languages. (The words ijtihad and jihad are nouns derived from these two verbs.)Īustronesian and Austroasiatic languages An example is اجتهد ijtahada "he worked hard", from جهد jahada "he strove". It is placed after the first consonant of the root an epenthetic i- prefix is also added, since words cannot begin with a consonant cluster. Arabic Īrabic uses a common infix, ⟨t⟩ ت for Form VIII verbs, usually a reflexive of Form I. standard Oscarito) Edgar → Edguítar Victor → Victítor. ![]() ![]() In Nicaraguan, Costa Rican, and Honduran Spanish, the Spanish diminutive affix becomes an infix ⟨it⟩ in names: Óscar → Osquítar (cf. Ancient Greek la mbánō (also with -an- suffix) "I take" (cf.perfect passive participle victus "conquered") Sanskrit exhibits the greatest transparency of this feature amongst the Indo-European languages, with the phenomenon manifesting in three of the ten traditional verb classes, where the infix is higher-grade and accent-bearing in the strong forms, and reduced-grade in the weak forms.This phenomenon is inherited, and preserved to varying degrees, by some early daughter languages such as Sanskrit, Ancient Greek, Latin language, etc. The stems of the other tenses have the root without the infix, and thus these verbs are called nasal-presents. The present tense of some Proto-Indo-European verbs, in the case of a certain number of roots, adds a nasal infix ( m, n) to the basic root. The use of ' expletive infixes' such as -fucking- and -bloody-, which are words rather than affixes, is known as tmesis.Infixes also occur in some language games.(education) This exists as a slang phenomenon. The ⟨-ma-⟩ infix (or "Homeric infix," after Homer Simpson), gives a word an ironic pseudo-sophistication, as in sophisti -ma-cated (sophisticated), saxo -ma-phone, (saxophone) and edu -ma-cation.The infix ⟨-iz-⟩ or ⟨-izn-⟩ is characteristic of hip-hop slang, for example h -iz-ouse for house and sh -izn-it for shit.None of the following are recognized in standard English. Most are heard in colloquial speech although there are other examples, such as in technical terminology, these examples are often more accurately described as tmesis. English Įnglish has almost no true infixes and those it does have are marginal. 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It provides an easy way to update maps, register products, and transfer data to and from Garmin devices. Garmin Express is a powerful software application that allows users to manage their Garmin devices. ![]() We use this information to complete transactions, fulfill orders, communicate with individuals placing orders or visiting the online store, and for related purposes. Online Storeįor orders and purchases placed through our online store on this site, we collect order details, name, institution name and address (if applicable), email address, phone number, shipping and billing addresses, credit/debit card information, shipping options and any instructions. We use this information to address the inquiry and respond to the question. 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To add video to the DVD, click the Movies button in the Media pane, then drag one or more movies into the drop boxes. (To access additional themes, use the pop-up menu above the row of thumbnails.) Choose a theme for the DVD by clicking one of the theme thumbnails. Use Magic iDVD when you want navigation menus and more than one piece of content on your DVD-for example, a couple of movies and some slide shows. As page 267 describes, OneStep DVD creates an autoplay, or kiosk-mode, DVD: the disc begins playback as soon as you insert it into a computer or DVD player. When you use OneStep DVD, you don’t have the opportunity to customize menu designs-there aren’t any. Use OneStep DVD when you want to burn just one movie to a DVD and you don’t need navigation menus. IDVD provides two ways to go from zero to DVD with very few steps. Indeed, Magic iDVD is a great way to rough out a project that you plan to refine later. And if you need to customize or enhance the DVD it creates, you can bring the rest of iDVD’s authoring features to bear. Magic iDVD may be all you need for many projects. Then click the Burn or Create Project buttons, and iDVD builds your project for you, even creating chapter submenus for movies containing DVD chapters (page 259). When you’re done, preview your work by clicking the Preview button and using iDVD’s standard preview features (page 255). Want a music soundtrack for a slide show? Drag an audio track into the slide show’s drop box. Drag an entire event or album from iPhoto or build a slide show one photo at a time by dragging individual photos into the same box. To create DVD slide shows, drag photos to the drop boxes. ![]() Magic iDVD presents you with a single window containing a list of menu themes, a set of drop boxes for holding movies and photos, and a media browser for accessing your audio, photos, and movies.Ĭhoose a theme, then drag movies into the drop boxes. If you want to burn a DVD to DVD using iDVD, scroll down to study the iDVD tutorial on how to import a DVD for latering burning into a blank DVD disc.When you want to create a DVD in a hurry, use the Magic iDVD feature. The DVD burning speed depends on the your computer performance, the video size. ![]() And it will start to burn projects from iMovie, iTunes library, hard disk etc. The recommended type is DVD-R that is playable on most DVD players for Mac.Ĭlick the Burn button, insert a blank disc into your computer's SuperDrive for Mac or Mac external hard drive - if your hard drive is new, learn how to easily format hard drive on Mac before using. Preview your projects to make sure they are ready before you continue to learn how to use iDVD to burn a DVD. Step 10: Burn Video to DVD on macOSĪfter the editing is finished, you can burn these projects to a DVD disc for backup or playing on TV, computer with disc drive. Choose Edit on the main menu > Select All buttons and edit button text, shape, image and add transitions to buttons. ![]() After a recent update, the entire library is available in lossless hi-fi at no extra charge. Pros: Apple Music has a library of 90 million songs that can be accessed on Apple, Windows, and Android devices. Who it’s best for: People who already have large iTunes libraries or who are otherwise committed to the Apple ecosystem. You don’t get to use the app any other way. That lets you access any song or playlist using your voice-and only your voice. You can use Siri to control Apple Music with a regular plan, but there’s also a cheaper “ voice plan” for $5 per month. There’s no free tier, but you can get a one-month free trial or six months free if you’ve bought eligible AirPods, HomePod smart speakers, or Beats headphones. Apple Music also has a discounted rate of $5 for students. Price: Individuals pay $10 per month for families of up to six, the service costs $15. And while Prime members don’t have to pay extra for the basic tier of Prime Music, its 2 million songs amount to a thin selection compared with what you get from other streaming services. The service also has a large and growing library of spatial audio content mastered in Dolby Atmos and 360 Reality Audio.Ĭons: Amazon Music Unlimited doesn’t seem to be quite as effective in making recommendations as many competitors, Spotify in particular. That entire library can now be streamed in high definition, and over 7 million tracks are available in 24-bit Ultra HD. Unlimited has more than 90 million songs, curated playlists, podcasts, and personalized stations. Pros: Both are ad-free, on-demand services, and if you already have a Prime subscription, Amazon Music Unlimited is the best deal you can get for a really robust collection of content (though not by much). ![]() And this is a solid option for streaming high-quality files. The deal isn’t as sweet as it used to be, but a lot of other services cost 99 cents more. Meanwhile, Amazon Music Unlimited is a good bet for Prime members looking for a bargain. Who it’s best for: Amazon Prime Music doesn’t cost anything beyond your regular Amazon Prime membership, so the appeal is obvious. You can get a special $5 rate if you sign up for the Single Device Plan via an eligible Amazon Echo device.Īnyone can access free, ad-supported playlists and stations by asking Amazon’s Alexa digital assistant to play music. There’s a free three-month trial and a discounted family plan. It’s $10 per month for non-Prime members. It has a library of 2 million songs.Īmazon Music Unlimited, with a far bigger library of songs and more features, costs $9 per month for Prime members (or $89 for an annual subscription). Price: Prime Music is included free with Amazon Prime, Amazon’s paid subscription service that costs $15 per month or $139 per year. ![]() Monster Jam which is specially known for Sports, Motorsports/Racing.Prices & Competitors are subject to change. Please monitor the venue website for health and safety policies, including face covering and entry requirements, which are subject to change. has the largest inventory of Lucas Oil Stadium tickets, so keep checking for great deals on resale tickets. The Lucas Oil Stadium 2024/2023 event schedule includes Taylor Swift, Morgan Wallen and Las Vegas Raiders. All guests age 2 & older are required to have a ticket. There is also a carved-out space of a mere 200,000 sq. The Official Monster Truck Series only at Monster Jam. The Series Champion receives an automatic bid to the prestigious Monster Jam World Finals to compete for the title of World Champion. Engineered to perfection, these 12,000 pound monster trucks, including the legendary Grave Digger, Max-D, Megalodon, and more push all limits in Freestyle, Skills and Racing competitions. Fan health and safety remains a top priority for Monster Jam and our venue partners. Early Entry Pass serves as your Pit Party Pass from 1:30pm to 5:30pm. Get in an hour earlier and beat the crowds so you have more time to see the trucks and drivers. All fans age 2 and older are required to have a ticket. Enjoy an exclusive extra hour with the Pit Party Early Access Pass. Witness heated rivalries, high-flying stunts and fierce head-to-head battles for the Event Championship. 10:00 AM Ticket Office Opens 1:30 PM Gates Open 3:00 PM Start Time EVENT INFORMATION This event will follow the NFL Clear Bag Policy. ![]() Monster Jam is an action-packed motorsport with world-class athletes locked in intense competitions of speed and skill. All guests age 2 & older are required to have a ticket. |
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