New Publication by Allele Biotech Researchers on Adipogenesis

Title:

A Novel pro-adipogenesis factor abundant in adipose tissues and over-expressed in obesity acts upstream of PPARg and C/EBPa

Authors:

Yuhui Ni, Chenbo Ji, Bin Wang, Jie Qiu, Jiwu Wang, Xirong Guo

Abstract:

An important question about adipogenesis is how master adipogenesis factors (defined as being able to initiate adipogenesis when expressed alone) peroxisome proliferator-activated receptor (PPAR) initiate adipogenesis only in differentiating preadipocytes. The objective of our research was to find previously unidentified factors that are unique or highly enriched in cells of the adipocyte lineage during adipogenesis that may provide functional tissue specificity to preadipocytes. We reasoned that such factors may alter expression profile specifically in obese individuals. Omental adipose tissues were obtained from obese and non-obese male patients undergoing emergency abdominal surgery. mRNAs extracted from either group were used for suppression subtraction hybridization (SSH). Genes corresponding to mRNAs enriched in obese versus non-obese patients were identified through sequencing and further analyzed for tissue distribution. Out of ~20 genes, we found several that showed clear fat cell specific expression patterns. In this study, we functionally studied one of these genes, previously designated as open reading frame C10orf116. Our data demonstrated that C10orf116 is highly expressed in adipose tissue and is localized primarily within the nucleus. Over-expression studies in 3T3-L1 cells indicated that it up-regulates the levels of CCAAT/enhancer binding protein a (C/EBPa) and PPARg and promotes adipogenic differentiation starting from the early stage of adipogenesis. Over-expressed in omental tissues from obese patients, C10orf16 manifested the characteristics of an adipocyte lineage-specific nuclear factor that can modulate the master adipogenesis transcription factors early during differentiation. Further studies of this factor should help reveal tissue-specific events leading to fat cell development at the transcriptional level.

Link to the original publication: http://link.springer.com/article/10.1007%2Fs10863-012-9492-6
Here is Allele Biotech’s webpage showing relevant cell and development biology products: www.allelebiotech.com

American CryoStem Corporation (OTCQB:CRYO), announced the launch of its newest adult stem cell and adipose tissue collection center in Bellevue, Washington

From AlleleBlog: http://blog.allelebiotech.com/2012/12/american-cryostem-corporation-otcqbcryo-announced-the-launch-of-its-newest-adult-stem-cell-and-adipose-tissue-collection-center-in-bellevue-washington/

A public company doing business of preparing and providing adipose (fat) tissue and adipose derived adult stem cells, American CryoStem Corporation (OTCQB:CRYO), announced the launch of its newest adult stem cell and adipose tissue collection center in Bellevue, Washington. Dr. Fredric Stern will officially launch the new Stern Center Stem Cell Collection Service as the first to provide Adult Stem Cell and Tissue Banking services to the general public in the Seattle, Washington area.

“Having successfully worked with American CryoStem in the past we are truly excited about the official launch of these adipose tissue based services to the general public in Washington. I look forward to working with American CryoStem on educating my patients about the Regenerative Medicine benefits of “bio-banking” and the latest fat transfer cosmetic services now available at the center. I chose to affiliate my practice with American Cryostem because of their thorough scientific approach to stem cell banking and strict adherence to aseptic technique and FDA guidelines,” said Dr. Fredric Stern, the founder of The Stern Center and a plastic surgeon.

John S. Arnone, CEO said, according to a company news release, “We are excited to have a surgeon with Dr. Stern’s abilities and reputation associated with American CryoStem in the Seattle, WA area and look forward to a productive relationship with the entire Stern Center team. We remain committed to our “Gold Standard” clinical laboratory processing and storage reputation and strive to provide the best physician and patient services in the U.S. The newest stem cell collection center in our network represents our commitment to associate with leading physicians in the Regenerative Medicine Industry.”

Mesenchymal stem cells (MSCs) are typically the products of adipose tissue-isolated stem cells for regenerative medicine or, in this case cosmetic surgeries. The mesenchymal stem cells can also be isolated from bone marrow or embryos. They secret hormones once introduced into human bodies and help balance cytokines in the blood. It is reported that MSCs help reduce several disease symptoms and, in some countries, are used as “youth fountains” in anti-aging treatment. MSCs can be produced fairly easily, in our hands at least, from induced pluripotent stem cells (iPSCs). iPSCs, like embryonic stem cells, can be expanded indefinitely, differentiated into MSCs and all other cell types, and are being tested in various cell therapies including those that are mediated through the MSC stage.

NIDA Branch Chief, Jonathan D. Pollock, Ph.D., Encourages SBIR/STTR Grants on Reagent Kits Including iPSC

 From AlleleBlog:

“We’re interested in areas of genetics, in terms of smoking cessation, pharmacogenomics, treatment of substance abuse, and particularly right now, issues related to prescription substance abuse,” Jonathan D. Pollock, Ph.D., chief of the Genetics and Molecular Neurobiology Research Branch at NIDA’s Division of Basic Neuroscience and Behavioral Research, told GEN.

In addition to that solicitation, Dr. Pollock said, the branch is interested in supporting commercialization and development of products, resources, and services through SBIR/STTR relevant to brain research. They include protein capture reagents, proteomics, genomics, pharmacogenomics, molecular diagnostics, nanotechnology, gene delivery and viral vectors, identification of RNA and DNA sequences in formalin fixed nervous tissue, shRNA, microfluidics, epigenetics diagnostics, therapeutics, and tools to detect epigenetic modifications.

The branch is also looking to support commercialization and development of biomarkers, optogenetics, reagents for iPS and neural stem cells, technologies to uniquely barcode cell types, improved super resolution microscopy methods, in vivo gene expression imaging, automated sectioning, image acquisition and 3D reconstruction of electron micrographic sections, genetically encoded markers for electron microscopy, and “big” genomic and proteomic data, including data visualization, data contextualization, and data analysis.
“What we’re really looking for is products that you could basically commercialize coming out of research. These can be things that are either products or services. I think that there are opportunities, particularly for groups of individuals that have an idea, IP, and want to have a startup company.”
SBIR/STTR grants account for 2.8% of NIDA’s roughly $1 billion annual budget. NIDA spent $26.679 million on SBIR and STTR in fiscal year 2012, which ended September 30—up from $26.497 million in FY 2011. The number of SBIR/STTR research projects grants rose to 56 in FY 2012 from 44 a year earlier, according to the GEN article.

Allele Biotech’s CEO, Jiwu Wang, Ph.D., has worked with Dr. Pollock on a previous, VHH nanobody-related project under the NIDA SBIR program. He has just submitted a SBIR grant application based on Allele’s recently published mRNA-based reprogramming technology, after discussion with Dr. Pollock.

莫言不言则已，一言就无耻

木石64 于 2012-12-7 13:22:55 发布在 凯迪社区 > 猫眼看人

 

 百姓一万倍痛感黑夜来临——海子《秋日黄昏》1987年。

    据说，现在允许“异质思维”了，不知此“质”为何“质”？我们大家拭目以待

    ——此为时论的前语。

  

    莫言领奖，再一次成为焦点，在接受境外记者采访时说，“如果说一个作家在一种完全自由的状态下必定能够写出伟大的作品，那一定是假话，如果说一个作家在不自由甚至不太自由的环境下必定写不出伟大的作品，那也是假话。”

    这话表面看起来完全正确，逻辑上毫无漏洞，但稍有脑子的人，就可以看出，其实质混账到极点！无耻到极点！只有独裁体制下甘心情愿做高级奴才的文化人，才说得出这样表面正确实质无耻到极点的话。

    按 照莫言的逻辑，我们可以推导出同样正确的一系列混账话。比如这一句：“如果说莫言不是他父母的儿子，那一定是假话；如果说莫言户口本上写着是他父母的儿 子，就百分之百绝不可能是别人的儿子，那也是假话。”——此言当然是有人身攻击之嫌的混账话，但从逻辑上而言，它和莫言的话，是同样正确的。

    再比如这一句：“如果说希特勒是好人，那肯定是假话；如果说希特勒从来没有做过一丁点好事，那同样是假话。”

    再比如这一句：“如果说薄熙来老婆杀人薄熙来没有责任，那肯定是假话。可如果说薄熙来老婆杀人就要薄熙来负责，那同样是假话。”

    类似的正确言论可以不间断写下去。可是，如果你是个脑浆尚存的百姓，不妨想一想，说这种话的人，动机究竟何在？目的究竟何在？

    莫言为何敢公然说出如此无耻的言论？原因不外是三点：

    第一、他知道这话主子听了特别高兴，而他的主子依然权势熏天。也或者说，他知道自己获了这个奖以后，实际已经上升到主子地位，必须为千万尚未升上去的文人准文人作出示范和训导——你只管自己去好好写作，不要管自由不自由的事，自由不自由不重要！

    第二、他自以为自己得了诺贝尔奖，现在放个屁都有人说好——至少主子开办的无数报刊确实一定会轰然叫好。而骂他的人，现在已经不在莫言眼里了，那基本都是无足轻重的屁民——哪怕是有文化的屁民。

    第三、他确信绝大多数中国老百姓头脑里脑浆不多，而且多数已经成了浆糊状，看不出他话里的无耻内涵。

    如莫言一类的无数中国无耻文人，比如含泪请求、做鬼也幸福、四千万身家、中国创造了世界上最好模式等等、等等言论的专利人，已经成为中国的真正耻辱。独裁不是耻辱，而是不幸。而人格沦丧，道德堕落，廉耻全无，才是民族真正的奇耻大辱。 

    要使中国真正站起来，必须让中国人首先在人格上站起来，否则，经济再发达，“其抵屁”增长再迅猛，也只不过用来扩充解决问题人民的维稳队伍，用来增高压迫民众的人为大山，用来奖励高唱红歌的无耻文人，用来装饰华丽的遮羞墙而已！
 

Non-business repost by Allele outpost.

Allele Biotech Announces Opening of New Facility in San Diego

From AlleleBlog http://blog.allelebiotech.com/2012/11/allele-biotech-announces-opening-of-new-facility-in-san-diego/

Firsr Floor Multipurpose Lab

We’re happy to announce that after a long construction and moving process, Allele Biotech has passed its final inspection hurdle and is now officially open for business at our new location, a two-story building we recently purchased on Nancy Ridge Drive. This facility is located at the heart of the Sorrento Mesa biotechnology cluster and contains two floors of brand new lab space to support production and R&D, plus plenty of room for expansion. We’re also welcoming two tenants, Nano CELLect and MesaTech, as well as the non-profit research institute we helped launch earlier this year, the Scintillon Institute (www.scintillon.org).

For our local colleagues and valued customers, please feel free to contact us to schedule a personal tour of our new facility and to meet with our esteemed group of experts who are always willing to discuss in person, your scientific needs

Keep an eye out on this blog for important dates, promotions and more news and photos from the new facility as we prepare for our upcoming open house! Hope to see many of you then if not sooner!

New Product Notes: iPS reprogramming and transdifferentiation product line including the Potent 6F mRNA Reprogramming Premix, IVT templates for Ascl1, Myt1l and neuro D2 mRNAs.

Allele Biotech Building Front

The 2012 Nobel Prize for Physiology or Medicine is Awarded to Cell Reprogramming Scientists

Monday Sir John B. Gurdon and Shinya Yamanaka shared this year’s Nobel Prize for physiology or medicine for work that revolutionized the understanding of how cells and organisms develop.

“By reprogramming human cells, scientists have created new opportunities to study diseases and develop methods for diagnosis and therapy.”

This is the 3rd time that a Nobel Prize is awarded on a technology that we chose as our area of research and made contributions to the field. The other two are RNA interference (2005) and fluorescent proteins (2008).

2012 Nobel Prize, adult stem cell banking, disease model, iPSC, John B. Gurdon and Shinya Yamanaka, regenerative medicine

Allele Biotechnology Announces New advance in production of human stem cells

Allele Biotech News Release:

This week in the journal Scientific Reports (Nature Publishing Group) scientists from Allele Biotechnology describe an important advance in the generation of stem cells capable of producing all the different tissues of the human body. In an article entitled “Feeder-Free Derivation of Human Induced Pluripotent Stem Cells with Messenger RNA,” Allele’s scientists present the fastest and safest method yet for converting ordinary human skin cells into “induced pluripotent stem cells” (iPSCs).

The scientific efforts were led by Dr. Luigi Warren, whose pioneering work on “footprint-free” reprogramming using messenger RNA was the foundation for Allele’s breakthrough. Through the united efforts of Dr. Warren and the scientists at Allele Biotechnology, his technique was re-engineered to increase cell conversion efficiency and eliminate any use of potentially unsafe reagents, while substantially reducing the time and effort needed to make stem cells. Dr. Warren believes that because of its advantages this technology “should become the method of choice for iPSC cell banking.”

According to Dr. Jiwu Wang, corresponding author on the paper and CEO of Allele Biotechnology, “This advance in stem cell derivation will enable both fundamental scientific research and clinical applications which has been the mission of Allele Biotechnology from its inception.”

Allele Biotechnology and Pharmaceuticals Inc. is a San Diego-based biotechnology company that was established in 1999 by Dr. Jiwu Wang and colleagues. A research based company specializing in the fields of RNAi, stem cells, viral expression, camelid antibodies and fluorescent proteins; Allele Biotechnology has always striven to offer products and services at the cutting edge of research.

Allele Biotechnology and Pharmaceuticals Inc.
Jiwu Wang, Ph.D., 858-587-6645 Ext 3
President and CEO
iPS@allelebiotech.com
fax: 858-587-6692
www.allelebiotech.com
Press release by BusinessWire. Also see Yahoo!News, Reuters, The Herald, etc.

Choosing the Right Fluorescent Protein

From AlleleBlog:  http://blog.allelebiotech.com/2012/08/choosing-the-right-fluorescent-protein/

In 1994 the green fluorescent protein cloned from Aequorea victoria became the first in a long line of genetically encoded labels. Since that time, the fluorescent protein palette has expanded to cover the entire visual spectrum. With so many color variations and options, which fluorescent protein (FP) is best for your research? Three key factors are among the most important to consider: brightness, photostability, and aggregation.
Brightness is the most obvious factor that most researchers consider when choosing an FP. In general, the brighter the FP, the better it will perform under almost all experimental conditions. When evaluating an FP’s brightness, make sure to look at the critical optical parameters — extinction coefficient and quantum yield. The product of these two values for different FPs can be used to directly compare their brightness. Brighter FPs will have lower detection limits (i.e. the concentration at which the FP becomes visible above autofluorescence of other cell components), and will allow imaging with lower excitation light intensity, minimizing the possibility of phototoxic effects.
Photostability has increasingly become a consideration when researchers choose fluorescent proteins. Many FPs, even if they are initially quite bright, will photobleach under continuous excitation during imaging. In order to perform long-term imaging experiments or to do quantitative analysis, an FP with high photostability should be the first choice. Unfortunately, methods for measuring and reporting photostability vary widely in the scientific literature, so be sure to understand how your FP’s photostability was measured before trying to make comparisons!
Aggregation (or oligomerization) has been one of the major issues tackled in the development of FPs. Many wild-type FPs form tetramers, which aggregate badly when expressed as fusion tags in cells. Engineered monomeric forms of many FPs are now available, and these monomeric FPs should always be used when making fusion constructs. For simple expression markers, however, oligomerization is not usually a major concern, and the brightest possible FP should be used in this case.
As with other research tools, doing your homework and reading the primary literature is always the best approach to choosing the right FP for your project!

Cord Banking and iPS Cells

Is it time to do it? 
From AlleleBlog: http://blog.allelebiotech.com/2012/08/cord-banking-and-ips-cells/

Umbilical Cord Banking (UCB) has been a popular discussion topic in the United States since the first Cord Bank was established in New York in 1992. Since the first cord blood transplantation in 1988, there have been over 780,000 UCB donations to private banks and 400,000 UCB donations to public blood banks worldwide. There has been such a great number of donations because UCB is full of hematopoietic progenitor cells, which makes it a more desirable solution to genetic, metabolic and immune disorders, over bone marrow and blood. Because of the nature of UCB, the recipient does not need to be an immunological match, there is a lower rate of infection and it is much easier to acquire than bone marrow, making it the ideal form of treatment for many patients and practitioners.
Over twenty years later, a new technology is emerging that could provide some clarity to the “to donate or not to donate” debate: induced pluripotent stem cells (iPSC). Derived from adult cells, iPSCs have the potential to be used like UCB or reprogrammed into specific tissue like myocytes. This potential opens up banking to countless individuals born before 1992, who never had an option to bank their UCB. With this unbounded potential, should iPSCs be banked liked UCB? Supporters argue that there has been enough evidence thus far to start a bank, however, most people seem to agree that too much is unknown about iPSCs and their use in humans. With that, most are in agreement that iPSC research is absolutely needed so banking can become a reality in the future.
For now iPSCs will remain in the testing and research phase, however, based on current research, iPSCs have the potential to enhance Cord Blood that has already been banked, perhaps providing some relief to public banks in the future (Broxmeyer, 2010). Though the potential of iPSCs is endless, more work has to be done before they are placed in humans and considered a viable banking system.

Social Media for the Consumer

AlleleBlog http://blog.allelebiotech.com/2012/08/social-media-for-the-consumer/:

Business has become an ever changing, dynamic landscape where the power and information no longer lies strictly in the hands of “the right people.” In less than a decade Facebook has managed to link nearly a billion people from tech savvy teens to curious senior citizens. Similarly, Twitter sees over 340 million tweets that reaches over 500 million active users per day. Social media is a wide reaching technology with infinite possibilities that has been synthesized and is fueled by people, not corporations.
In a recent Industry report, 93% of businesses reported using social media as a marketing tool with the benefits being increased exposure and customer interaction (Stelzner, 2011). What does this study mean to consumers? Businesses are coming to you and responding to your feedback. Social media gives you, the consumer; back the power and the opportunity to benefit from your loyalty. Most businesses, from restaurants to labs, offer weekly tips or discounts on their services to the customers who “follow” or “like” them. Here at Allele Biotech, we have begun offering special weekly promotions, information about our products and interesting biotech news to our customers who either “follow” us on Twitter or “like” us on Facebook. For the entire month of August we are offering 15% off just for a “like” or “follow.” If you are one of the billion users, it doesn’t take much more than a click for instant rewards.
We recognize that the reason we create this technology and improve is because of the consumers. Allele Biotech would like to thank you for your loyalty and encourage you to remain active within our social media to push us forward and to help us continuously strive for more. With the creation and exponential popularity of social media, the power and information as become available to the most important people, the consumers.

Twitter @Allele_Biotech
Facebook http://www.facebook.com/pages/Allele-Biotechnology-and-Pharmaceuticals-Inc/78331924957

VHH Nanobodies in Super-Resolution Imaging and More

From AlleleBlog home page:

From the large number of recent publications using GFP-Trap beads, it appears that GFP-Trap is on the way to becoming one the most popular tags for co-IP thanks to its unparalleled “cleanness” of precipitated protein bands and its quantitative binding capabilities. As described previously, the antibody conjugated on the GFP-Trap beads is a single-domain antigen binding module from camelid single-chain antibodies. Termed VHH, this domain is only ~12 kD and can fit into structures that other types of antibodies cannot. We have successfully created VHH antibodies against a number of neural factors as a research project for the NIDA/NIH.

VHH proteins are often called nanobodies as a result of their size (1.5 – 2.5nm) and binding affinity ( GFP-trap has a binding affinity of 0.59nM). In addition to their use for co-IP, VHH antibodies have proven themselves as a resilient tool for various other applications. Anti-GFP nanobodies, for example, are currently used to enhance the fluorescence of GFP (GFP-trap booster utilizes the same VHH binding antibody coupled to a fluorescent dye); others have used VHH antibodies that can insert into certain part of GFP to dim the fluorescence signal . More recently, Ries et al. published in Nature Methods that the anti-GFP nanobodies offered a simple and versatile method for super-resolution imaging (i.e. PALM)-previously super-resolution imaging requires photoconvertible fluorescent proteins (such as Eos, mClavGR2). With dye-conjugated nanobodies, generating fusions to these newer FPs is no longer needed, however, using the nanobody super-imaging method requires fixing and permeabilizing the cells.

When using anti-GFP VHH reagents you need to be aware that other fluorescent proteins can also be recognized, if they were derived from the avGFP (jellyfish GFP). Also, some GFPs are not recognized if they are from another species, or engineered such as our mWasabi. We are producing newer and brighter GFP/YFPs based on the lancelet YFP protein to offer alternative series that will not be cross-recognized by the GFP-Trap antibodies.

Picture Blog — Making mRNAs by In Vitro Transcription for Transgene Expression and R-iPSCs

R-iPS Cell FAQ 2:
What is the expected yield from the in vitro trancription (IVT) reactions?

Performed as described, you should recover around 40 ug RNA from each 40 uL IVT reaction.

R-iPS Cell FAQ 3:
How can the success of the RNA synthesis protocol be assessed?

Run 500 ng (5 uL) of the concentration-adjusted products on an E-gel to check for consistent product yield and relative product sizes, and to confirm the absence of secondary bands or smears.

萧雪慧：历史还在忍辱含垢——写在林昭遇难44周年忌日

44年前的今天，一个智慧、高洁的美丽女性被密杀和灭尸了。她就是林昭。这个于1954年以江苏省最高分靠进北大中文系新闻专业的女大学生不仅才华出众、 勤奋多思、对真理有着执著追求，而且坦诚率直，心口如一。然而恰恰是这些高贵禀赋使她最终招来杀身之祸。在一个没有法律巍然屹立，权力运作无规可寻，全凭 掌握最高权力者反复无常的意志支配的地方，其统治是不准人们提异议的，而统治的权威则是建立在人们普遍的愚蠢、盲从和怯懦之上的。但凡权力运作以及建立和 维系权威的方式呈现为上述状况之处，无论以多么堂皇的主义自我标榜，其内里是彻头彻尾的专制主义。专制主义与人的高贵禀赋天然敌对。林昭在劫难逃。
　　　　
　 　1949年以后确立的新政权在夺取和建立政权前曾一再作出实现“民主”“自由”的庄严承诺，人们因此而普遍抱有这种合理预期。但现实与承诺之间的巨大反 差使许多人在深深的失望中酝酿着强烈不满的思潮。尤其在尚保持着独立思想的知识界，种种不满和怀疑思潮有力地涌动着。这种情况对于其统治不准提异议的人来 说是危险的。一场主要针对知识界的“阳谋”就在这样的背景下付诸实施了。正是始自这场名曰“引蛇出洞”的“阳谋”而对思想和文化的史无前例的大屠杀开始了 林昭的厄运。11年后，她又在另一场对思想和文化更加史无前例的大屠杀中被从肉体上消灭了，而惨无人道的残杀和灭尸手段暴露出置她于死地的一方对她的深仇 大恨。这是暴政下通行的“小人政治”对精神高贵者的深仇大恨。
　　　　
　　在1957年那场凭借国家暴力实施的“阳谋”中，林昭起初并 不在因言罹祸的那批学生之列。她的厄运源自她在良心和识时务之间义无返顾地选择了良心。尽管大学生群体在这场向知识界袭来的风暴中迅速崩溃瓦解，面对残酷 现实，各人有着很不相同的选择：有人沉默；有人为了自保而表违心之态，与落水者拉开距离甚至划清界限；更有人从包围着知识界的暴力氛围中嗅出机遇，选择了 以告密、扣帽、打落水狗等卑劣下贱行径为进身之阶......。在各色人等作各式表演时，并未入另册的林昭如果保持沉默，她完全可以以不失体面的方式保全 自己。可林昭天性高贵，不愿把自保置于信念和尊严之上。她在风暴中既没有停止危险的思想求索，也不屑因为怕湿脚而沉默。在一场早已受到监控的辩论会上，她 毅然站出来为处于一大帮自称“左派”的狙击手火力焦点下的同学仗义执言：“不是号召党外的人提意见吗？人家不提，还要一次一次地动员人家提！人家真提了， 怎么又勃然大怒了呢？”她坦陈自整风以来一直没有说话、没有写什么，是料到“一旦说话也就会遭到像今晚这样的讨伐”；坦陈曾经困扰着她的“组织性与良心的 矛盾”。对一个发自阴暗角落的威胁性质问“你是谁？”她磊落地以“‘双木三十六’之‘林’，‘刀在口上之日’的‘昭’”朗朗作答。
　　　　
　 　就这样，林昭因为选择了良心、因为面对威吓表现了磊落和不屈而当了“右派”，其后又因为不屈不挠的求索精神和对被许多人弃之若敝履在她却是安身立命之本 的人的尊严、人的思想言论自由的维护而遭到惩罚，暴政施于她的残害步步升级：从57年被打入另册到68年被杀害的这11年中，对付她的措施由监管改劳教， 由劳教改二十年重刑，直至干脆从肉体上消灭。每一次为残害升级而罗织的罪名十分荒唐—— “抗拒改造”、“不认罪”、 “态度恶劣”，——这是一些在任何类型国家的刑法中找不到的罪名。然而对于行暴政的人来说，这是最不可饶恕的。暴政统治靠把人愚化、劣化和侏儒化来维系， 只有把所有人变成精神白痴，只有打断所有人的精神脊梁、摧毁所有人的自尊，只有把社会环境败坏成高尚者的地狱、卑劣小人的乐园，它才是安全的。林昭身上有 着最令暴政统治仇恨和不安的精神气质，这就是与适应暴政需要的精神愚化、劣化、侏儒化截然对立的智慧、良知和傲然挺立的人格尊严。当人们在有组织的和系统 的谎言包围中、在精神暴力和物质暴力的威逼下普遍为自保而选择了放弃思想和言说的权利、普遍变得愚蠢甚至选择了愚蠢时，她既不愿把大脑弃置不用，也不愿沉 默。她不顾自身处境、蔑视种种思想禁令而不断求索。她的智慧和探索精神使她发现了许多被刻意掩盖着的真相、被有组织的和系统的谎言所扭曲甚至颠倒的事实， 如彭德怀事件、“大跃进”与人民的苦难、南斯拉夫问题......等等，这些于八十年代初的思想解放运动中才开始逐渐揭开真相的问题，她早在1960年就 以自己的独立思考接近了事实真相，她理当属于我们这个苦难民族的先知先觉者，当然也成了最遭暴政忌恨的目标——尤其是她不肯放弃说出已经被她窥破的真相。 因为这，对她的处理从劳教升级为二十年重刑。被投入监狱的林昭不向强权低头，她只身撞击黑暗，在惨无人道的摧残下以自己的柔弱之躯承当起对思想言论自由这 些源自人的类本质的神圣权利的坚决捍卫，既不掩饰自己的观点和信念，更不掩饰对强权的蔑视。坚守智慧、良知、人格尊严——这就是“抗拒改造”！这就是“不 认罪”！这就是“态度恶劣”！它们击中的是暴政在强大的表象背后脆弱的根基，所以罪不可赦。林昭在经历了暗无天日的八年狱中生活摧残后因这强权政治下的不 赦之罪被秘密杀害。她是在口被堵住的情况下遇害的。遇害后，她的血肉之躯遭到凶手毁尸灭迹；记载了她的思想、信念和情感世界的诗文、书信、日记等一切有形 的文字材料、照片、甚至包括她在狱中用纸折叠的工艺品等一切物件也被销毁了。采取这些非比寻常的野蛮手段，显然是根基脆弱的强权要消灭掉有这样一个人只身 挑战黑暗、撞击黑暗的任何痕迹。
　　　　
1852年，雨果针对路易.波拿巴政变期间发生在巴黎蒙马特大路的屠杀愤怒地写下了一段话： “历史是一位伟大的忍辱含垢者，它被迫接受这令其汗颜耳赤的史实”。一百多年后发生在中国大地上对思想、文化一场又一场的大屠杀，死难者不知多少万倍于巴 黎蒙马特大路的屠杀，我们的历史被迫接受的是更加令其汗颜耳赤的史实；无数在暴政残害下丧身的死难者中，林昭因精神高贵而罹难的全过程尤其使历史蒙羞。如 今，四十多年过去了，暴政强加给历史的耻辱并未洗刷，甚而既因为有组织的掩盖、伪造历史而加深，也由于邀宠者把那个曾经吞噬了无数人的时代指鹿为马说成 “阳光灿烂的日子”而加深。1966年，林昭的亲人最后一次在狱中与她相见，知道自己随时会被杀的她留下一个遗愿：“相信历史总有一天人们会说到今天的苦 难！希望把今天的苦难告诉给未来的人们！”林昭遇害是在两年后。而她留下这个遗愿的时间距今已经46年——快半个世纪了。然而直面历史、不遮不掩地说出当 年苦难的日子至今还未到来，令林昭最为痛恨的精神欺骗也还在继续，历史还在被迫忍辱含垢。在林昭遇害44周年忌日，我愿以这篇短文作为对林昭遗愿的回应， 并愿与所有希望洗刷历史耻辱的人们一道为此而做一些应做之事。

陈光诚自由！ Chen Guangcheng is free and likely under US protection in Beijing Embassy

难以相信一个盲人，在没有别人帮助下能自己计划几个月，利用24监视的疏忽和漏洞，选择月亮暗淡的晚上，用十几秒的时间翻墙越“狱”，在溪流旷野间奔跑几小时，摔倒几十次，知道谁是可靠的网友，联系上并立即转移城市，录制对总理温家宝的呼吁。CNN report called it “like a Hollywood thriller”, 但陈光诚的行动和语气中显示出来的决心，勇气，智慧， 和冷静实在是超越thriller movie所能描绘的英雄。

当然他不可能没有同样具有勇气和牺牲精神的朋友的帮助，何培蓉在Skype接受CNN采访时显示出来的镇定“I'm not concerned about my own safety," she said during the interview. "I hope they'll arrest me, not my friends.”, when the state security agents just arrived at the door，is legendary. 自己就曾经历过类似关押的胡佳显然在这次陈光诚行动中起了很大的作用。

Social concern blog by Allele Outpost

Picture Blog — Human mRNA-Induced Pluripotent Stem Cells Generated in Days

R-iPS Cell FAQ 1:
What phenotypic changes can be observed during a successful reprogramming trial?

About one week out, target fibroblasts should show an involution of fibroblastic processes, and foci or clusters of epitheliod cells—ideally with small nuclei, minimal cytoplasm, and signs of ongoing mitosis—should appear. Colonies with hESC morphology typically start emerging in ~10-14 days.

From AlleleBlog: http://blog.allelebiotech.com/2012/04/picture-blog-human-mrna-induced-pluripotent-stem-cells-generated-in-days/

Making brown adipose tissues from RiPSCs

From AlleleNews

Compared to the generation of other tissue types, adipogenesis is relatively well studied because of the long availability of mouse cell line 3T3-L1 that can be readily differentiated into white adipocytes upon a simple treatment of a few chemicals. For human adipogenesis cell models, either mesenchymal stem cells (MSCs) from bone marrow or other tissues or adipose-derived stromal vascular cells18 (ADSVCs) could be derived into fat cells. However, these cells have limited potential for expansion for continued studies.

Using iPS cells for tissue development has been a hot topic in many fields of developmental biology. Although one could transfect or transduce iPSCs, it is still often necessary to grow the stem cells into some type of progenitor cells through 3-D suspension culture as embryonic bodies. In a recent Nature Cell Biology paper, Ahfeldt and colleagues in Cowan lab created MSCs from RiPSCs generated by the Luigi Warren mRNA method, then derived them into either white adipose tissues (with lentivirus carried PPARG2) or brown (PPARG2–CEBPB–PRDM16) adipose tissues. It is particularly interesting that brown fat tissues, often considered the “good fat”, could be derived in a dish. This study will open doors for further use of the footprint-free mRNA-derived iPSCs for virtually unlimited supply of precursor cells for in vitro differentiation.

News and Views: http://www.nature.com/ncb/journal/v14/n2/full/ncb2430.html#/references

Original article: http://www.nature.com/ncb/journal/v14/n2/full/ncb2411.html

Opportunities for business with Allele Biotech

AlleleBlog: http://blog.allelebiotech.com/2012/02/opportunities-for-business-with-allele-biotech/

Allele Biotech is known for staying on the edge of biological research fronts when it comes to developing new technologies into useful tools. Our research also has far-reaching implications and potential applications outside of the traditional biomedical research reagent field. Some of these technologies were the results of researchers interacting with the Allele scientific team, who wanted Allele to help realize their potentials. If you are interested in investing, co-developing, or trading in our areas of expertise, please email us at oligo@allelebiotech.com.

1) A novel method of discriminating and/or detecting mismatched polynucleotide populations in a sample, or determining the relative abundance of the species contained in the sample based on the changes in the relative ratios following a critical treatment. This technology, subject of a current patent application, can provide great benefits in polynucleotide-based diagnosis.

2) A technology on how to utilize the light-absorbing capabilities of certain light-absorbing proteins against damaging lights, or in cosmetic or beauty products. It is also a subject of a filed full patent.

3) Products that relate to detecting swine flu with novel antibodies of high specificity and stability. The antibodies have been tested in academic molecular biology labs in ELISA and strip formats.

4) Nanotechnology products that can be immediately applied to prevent citrus diseases on farms.

Making Transfection-Grade mRNA by IVT (In Vitro Transcription)

RNases are an often feared in molecular biology labs because of their high stability and ominous presence in virtually all living systems. Consequently, people who work with RNA are trained to exercise extreme caution to avoid RNA degradation: change gloves often because human hands ooze RNases; use only sterilized labware as microbes may be sources of RNases; for surfaces that can’t be autoclaved, use sprays like “RNase Zap” (SDS- or guanidine-containing solutions). Such cautionary steps are especially necessary when dealing with low abundance RNA samples.

RNAs can be produced by in vitro transcription (IVT), a simple reaction requiring only a DNA template (double-stranded or even single-stranded DNA as long as the promoter region is double-stranded), RNA polymerase (from T7, SP6, or T3 phage), NTPs, and a reaction buffer that provides appropriate salt and pH. Standard NTPs may be replaced with modified ones to either increase stability or to reduce immune-response when transfected into cultured cells. Additionally, a 5’ cap structure may be added during IVT for further stabilizing mRNAs inside the cells post transfection. Using a commercially assembled kit, one can routinely produce 40-50 µg of mRNA from 1 µg of DNA template in a single 20-50 µl reaction.

At such high concentrations, IVT mRNAs are not nearly as sensitive to RNase-mediated degradation as low-abundance samples. The mRNA can be easily observed on agarose gels that are regularly used for DNA, and their integrity can be monitored after transcription or storage. In most cases one distinct band of mRNA from an IVT reaction is obtained as long as a clean DNA template is used. Preparing a good, uniform IVT template is critical to prevent aberrant products. By using high quality templates, IVT mRNA produced in your own lab are often higher in quality than mRNAs purchased from current commercial sources (Figure in Blog shows mRNAs generated by IVT for R-iPSC). Sometimes there are minor bands created during IVT, but they normally do not interfere with the intended uses of the mRNA, and can be purified away with a purification kit (by using a discriminating purification scheme such as Allele Biotech’s Surface Bind RNA Purification, smaller species can be specifically removed, a separate topic for another blog).

Once produced, mRNAs can be stored at -20C for months, or -80C nearly indefinitely.

http://blog.allelebiotech.com/2012/01/making-transfection-grade-mrna-by-ivt-in-vitro-transcription/