• 我要登录|
  • 免费注册
    |
  • 我的丁香通
    • 企业机构:
    • 成为企业机构
    • 个人用户:
    • 个人中心
  • 移动端
    移动端
丁香通 logo丁香实验_LOGO
搜实验

    大家都在搜

      大家都在搜

        0 人通过求购买到了急需的产品
        免费发布求购
        发布求购
        点赞
        收藏
        wx-share
        分享

        miRNA克隆中的问题,拜托高手了,急~~

        丁香园论坛

        1259
        Cloningof miRNAs was performed as described (Elbashir et al., 2001; Llave
        et al., 2002a). In brief, ;600 mg of RNA was resolved through two lanes
        on a denaturing 15% polyacrylamide gel. Labeled DNA oligonucleotides
        were used as size standards. A gel fragment spanning the size range
        of 15 to 26 nucleotides was excised, and RNA was eluted overnight with
        0.4 M NaCl at 4℃. The RNA was recovered by ethanol precipitation, dephosphorylated,
        and again ethanol precipitated. Then, the small RNAs
        were ligated sequentially to 5’ and 3’RNA/DNA chimeric oligonucleotide
        adapters (Dharmacon Research, Boulder, CO). The 3’ adapter oligonucleotide
        (5’-pUUUaaccgcatccttctcx-3’; uppercase, RNA; lowercase,
        DNA; p, phosphate; x, inverted deoxythymidine) (Elbashir et al., 2001)
        possessing a 5’ monophosphate and a 3’ inverted deoxythymidine to
        prevent self ligation, was then ligated to the dephosphorylated small RNA.
        The ligation product was recovered from the gel, 5’ phosphorylated, and
        the RNA was recovered by ethanol precipitation. Next, the 5’ adapter
        (5’tactaatacgactcactAAA; uppercase, RNA; lowercase, DNA) was ligated
        to the phosphorylated ligation product as described above.

        这是文章里克隆小RNA的方法,我是搞不懂他的3‘和5’街头是怎么设计的,他是用的DNA/RNA杂和的接头,可是给的序列只有一条??
        里面的uppercase是指上面的那条吧,谁能告诉我他的这个接头的设计原理,我知道里面有一个酶切位点!!
        谢谢

        yongdali@hotmail.com
        QQ:153206517
        ad image
        提问
        扫一扫
        丁香实验小程序二维码
        实验小助手
        丁香实验公众号二维码
        扫码领资料
        反馈
        TOP
        打开小程序