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Prepare manuscripts according to the Publication Manual of the American Psychological Association using the 7th edition. Manuscripts may be copyedited for bias-free language (see Chapter 5 of the Publication Manual). APA Style and Grammar Guidelines for the 7th edition are available.
We designed a modified STARR-seq reporter construct pGL4.10-Sasaki-SS (a) based on an earlier published design15 in the pGL4.10 backbone (Promega, E6651). The sequence between SacI and AfeI was replaced with a sequence containing CG-depleted chicken lens δ1-crystallin gene (Sasaki) promoter51, a synthetic intron (pIRESpuro3; Clontech, 631619), an ORF (fusion of Nanoluc-EmGFP), homology arms for library cloning with AgeI and SalI restriction enzyme (RE) sites flanking the ccdB gene, a small 52-bp DNA stuffer (a part of the neomycin resistance cassette) and a 20-bp sequence from the 3ʹ-Illumina adapter for optimally sized final library for Illumina sequencing and the SV40 late poly(A) signal from the pGL3 backbone (Promega, E1751).
To enable the analysis of CpG methylation on enhancer activity, we designed modified STARR-seq vectors in a CpG-free backbone with Lucia reporter gene (Invivogen, pcpgf-promlc) driven either by the EF1α promoter (b. pCpG-free-EF1α-SS) or the Sasaki promoter (c. pCpG-free-Sasaki-SS-v1) as above. To facilitate the cloning of the synthetic DNA library to the 3ʹ UTR of the reporter gene, the cloning cassette from the pGL4.10-Sasaki-SS vector (a) containing the homology arms with AgeI and SalI RE sites, the 52-bp DNA stuffer and the 20-bp sequence from the 3ʹ-Illumina adapter as above was introduced to the CpG-free vectors using the NheI site.
Despite the complexity of the virus and the industry wide slowdown at the end of the year, the business performed well during a difficult time and 2021 was another good year for our company. Total revenue grew greater than 20% and our adjusted EBITDA margin expanded more than 200 basis points. As we take stock of how far Chegg has come and the opportunity ahead of us, the past two years have been truly remarkable. Since 2019, Chegg Services revenue, adjusted EBITDA, and free cash flow have more than doubled. This all while investing in future growth initiatives such as Learn with Chegg, Uversity, Chegg Life, plus expanding into skills-based learning and into international markets. Additionally, through acquisition we have made key assets such as Mathway in 2020 and more recently our acquisition of Busuu, allowing us to expand into new subject matters and geographies.
Looking at the balance sheet, we ended the year with cash and investments of $2.3 billion. This was bolstered by the aforementioned free cash flow of $177 million, primarily offset by a $300 million accelerated share repurchase, or ASR, we entered into in early December and completed two weeks ago, retiring approximately 10.6 million shares. We entered into the ASR as we believe there was an overreaction to the temporary headwinds that affected our industry, which caused a dislocation in our share price. We believe this was a good use of our capital, good corporate hygiene and increases shareholder value. 2ff7e9595c
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